MicrobiologyBytes

Despite the availability of antibiotics that rapidly kill bacteria in vitro, the treatment of chronic bacterial infections, such as tuberculosis, requires long-term drug therapy. The reasons for this are unclear, but many have hypothesized that the slow replication and concomitantly low metabolic rate of bacteria in the host environment produce an “antibiotic-tolerant” state. Researchers tested this hypothesis by identifying the bacterial pathways responsible for slowing the growth and metabolism of Mycobacterium tuberculosis in response to stress. They found that diverse growth-limiting stresses trigger a common signal transduction pathway that slows bacterial growth by redirecting cellular carbon fluxes away from central metabolic pathways and towards storage. Disruption of this metabolic switch increased the antibiotic sensitivity of the bacterium during infection, verifying that this response significantly contributes to antibiotic tolerance and suggesting new strategies for accelerating therapy.

Metabolic Regulation of Mycobacterial Growth and Antibiotic Sensitivity. (2011) PLoS Biol 9(5): e1001065. doi:10.1371/journal.pbio.1001065
Treatment of chronic bacterial infections, such as tuberculosis (TB), requires a remarkably long course of therapy, despite the availability of drugs that are rapidly bacteriocidal in vitro. This observation has long been attributed to the presence of bacterial populations in the host that are “drug-tolerant” because of their slow replication and low rate of metabolism. However, both the physiologic state of these hypothetical drug-tolerant populations and the bacterial pathways that regulate growth and metabolism in vivo remain obscure. Here we demonstrate that diverse growth-limiting stresses trigger a common signal transduction pathway in Mycobacterium tuberculosis that leads to the induction of triglyceride synthesis. This pathway plays a causal role in reducing growth and antibiotic efficacy by redirecting cellular carbon fluxes away from the tricarboxylic acid cycle. Mutants in which this metabolic switch is disrupted are unable to arrest their growth in response to stress and remain sensitive to antibiotics during infection. Thus, this regulatory pathway contributes to antibiotic tolerance in vivo, and its modulation may represent a novel strategy for accelerating TB treatment.

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The Paramyxoviridae are enveloped, non-segmented, negative-strand RNA viruses that include major human pathogens belonging to two subfamilies. The Pneumonvirinae subfamily includes respiratory syncytial virus (RSV) and the metapneumoviruses, while the Paramyxovirinae subfamily includes, amongst others, measles virus (MeV), mumps virus, human parainfluenza viruses (hPIV1-4), and the recently emerged, highly pathogenic henipaviruses Hendra (HeV) and Nipah (NiV). Members of both subfamilies are responsible for significant human morbidity and mortality. MeV, in particular, remains a major cause of childhood mortality worldwide despite the availability of a live-attenuated vaccine.

All paramyxoviruses gain entry into and spread between cells by promoting direct membrane fusion. Membrane merger is mediated by the viral fusion (F) protein, which, like other class I fusion proteins such as influenza HA and HIV env, first forms metastable homo-trimers that require proteolytic activation to gain functionality. Receptor binding by the attachment protein is thought to then trigger major conformational changes in mature F, resulting first in insertion of a hydrophobic domain, the fusion peptide, into the target membrane and ultimately in formation of a fusion pore through juxtapositioning of the F transmembrane domain and fusion peptide in the thermodynamically stable postfusion conformation. Unlike retro- or orthomyxovirus entry, the complexity of the paramyxovirus fusion triggering mechanism is raised to a higher level by the fact that the receptor binding and fusion-promoting functions are contributed by separately encoded envelope glycoproteins. This physical separation of the two functions necessitates a mechanism of posttranslational linkage, which is accomplished through the formation of virus-specific hetero-oligomer complexes between the two proteins. However, the overall organization of functional Paramyxovirinae fusion complexes and the molecular mechanism that links receptor binding with coordinated F protein refolding into the postfusion conformation remain largely unknown.

Structural and Mechanistic Studies of Measles Virus Illuminate Paramyxovirus Entry. (2011) PLoS Pathog 7(6): e1002058. doi:10.1371/journal.ppat.1002058
Measles virus (MeV), a member of the paramyxovirus family of enveloped RNA viruses and one of the most infectious viral pathogens identified, accounts for major pediatric morbidity and mortality worldwide although coordinated efforts to achieve global measles control are in place. Target cell entry is mediated by two viral envelope glycoproteins, the attachment (H) and fusion (F) proteins, which form a complex that achieves merger of the envelope with target cell membranes. Despite continually expanding knowledge of the entry strategies employed by enveloped viruses, our molecular insight into the organization of functional paramyxovirus fusion complexes and the mechanisms by which the receptor binding by the attachment protein triggers the required conformational rearrangements of the fusion protein remain incomplete. Recently reported crystal structures of the MeV attachment protein in complex with its cellular receptors CD46 or SLAM and newly developed functional assays have now illuminated some of the fundamental principles that govern cell entry by this archetype member of the paramyxovirus family. Here, we review these advances in our molecular understanding of MeV entry in the context of diverse entry strategies employed by other members of the paramyxovirus family.

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Going somewhere exotic? If you are, make sure to look at the Malaria Hotspots website, full of essential information for travellers:

• Between 1990-2009, every year approximately 1,800 British travellers return home with malaria. The UK is one of the biggest importers of malaria in Europe.
• The most severe form of malaria (Plasmodium falciparum) accounted for 79% of cases amongst British travellers in 2009.
• Malaria is a preventable infection but can be fatal if left untreated – an average of nine people die each year from malaria in the UK.
• Malaria is transmitted by infected mosquitos. It only takes one bite from an infected mosquito to contract malaria.

The Malaria Hotspots website is an educational initiative organised and funded by GlaxoSmithKline Travel Health.

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On June 5 1981, MMWR published a report of Pneumocystis carinii pneumonia in five previously healthy young men in Los Angeles, California; two had died. This report later was acknowledged as the first published scientific account of what would become known as human immunodeficiency virus (HIV) and acquired immunodeficiency syndrome (AIDS). Thirty years after that first report, the most recent estimate is that 33.3 million persons were living with HIV infection worldwide at the end of 2009.

In 1981 I was working on my PhD (on poliovirus) when I first heard about AIDS. Someone from the department came back from a trip to San Francisco with lurid and scary tales. Over the next couple of years, my interest in retroviruses grew and when I finished my PhD in 1984, I went off to California to work on HTLV and HIV.

In the United States, CDC estimates that 1,178,350 persons were living with HIV at the end of 2008, with 594,496 having died from AIDS since 1981. At this 30-year mark, efforts are being accelerated under the National HIV/AIDS Strategy of the United States, with goals of reducing the number of persons who become infected with HIV, increasing access to care and optimizing health outcomes for persons living with HIV, and reducing HIV-related health disparities.

I worked on HIV for 10 years before moving on to other things. I still maintain a close interest in the topic. 30 years – it seems like yesterday.

MMWR 60(21): 689 June 3 2011

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Nanomedicine refers to the medical application of nanotechnology, and particularly to the development of novel nanomaterials that can be used for disease diagnosis and therapy. The unique properties of nanoparticles promise to deliver a new generation of diagnostic reagents with higher signal-to-noise ratios than current imaging modalities, as well as targeted therapies that are more efficacious than today’s medicines and that have fewer adverse effects. Nanomaterials have a large surface-to-volume ratio compared to traditional delivery vehicles that offers a greater capacity for drugs and/or imaging reagents, and the ability to decorate nanoparticles with specific ligands means these diagnostic and therapeutic payloads can be delivered to particular cells. Several classes of nanomaterials are currently being developed, including synthetic materials and naturally occurring bionanomaterials such as viral nanoparticles (VNPs). Each of these systems has benefits and limitations with regard to pharmacokinetics, toxicity, immunogenicity and specificity for the target tissue.

Will virus-based nanoparticles revolutionize medicine or will it all end in grey goo?

Applications of viral nanoparticles in medicine. Curr Opin Biotechnol. May 16 2011
Several nanoparticle platforms are currently being developed for applications in medicine, including both synthetic materials and naturally occurring bionanomaterials such as viral nanoparticles (VNPs) and their genome-free counterparts, virus-like particles (VLPs). A broad range of genetic and chemical engineering methods have been established that allow VNP/VLP formulations to carry large payloads of imaging reagents or drugs. Furthermore, targeted VNPs and VLPs can be generated by including peptide ligands on the particle surface. In this article, we highlight state-of-the-art virus engineering principles and discuss recent advances that bring potential biomedical applications a step closer. Viral nanotechnology has now come of age and it will not be long before these formulations assume a prominent role in the clinic.

E. coli has been in the media a lot recently (Latest News), so MicrobiologyBytes thinks it’s time for:

10 things you should know about E. coli:

1. Escherichia coli (E. coli) is a normal inhabitant of the human gut. It’s been with us for millions of years and overall does us a lot of good, e.g. helping with digestion and providing vitamins we can’t make for ourselves.

2. There are many different strains of E. coli, which all look much alike. They are identified by the antigens on the surface of the cell. These include somatic (O antigens) on the surface of the cell, flagellar (H antigen) and capsular (K antigens) associated with polysaccharide capsules on some strains.

3. A few strains of E. coli are pathogenic and cause disease. Enterotoxigenic (ETEC) strains cause diarrhea but are non-invasive and do not leave the intestine. Enteropathogenic (EPEC) strains also cause diarrhea and enter epithelial cells around the intestine. Enteroinvasive (EIEC) strains cause severe diarrhea and high fever. Enterohemorrhagic (EHEC) strains such as E. coli O157:H7 cause bloody diarrhea, hemolytic-uremic syndrome and kidney failure.

4. E. coli O157:H7 infections often case to bloody diarrhea and occasionally acute kidney failure, especially in young children and elderly people.

5. Most infections are associated with eating undercooked, contaminated ground beef (e.g. burgers), drinking unpasteurized milk, swimming in or drinking contaminated water, and eating contaminated salad vegetables. Infection can also be aquired via direct contact with animal faeces, for example on farms.

6. A bit of dirt never did me any harm… E. coli O157:H7 is new. It was first recognized around 25 years ago and is now widespread, possibly due to agricultural practices.

7. Where did it come from? This strain of E. coli contains lysogenic bacteriophages which encode Shiga toxins (these strains are known as STECs: Shiga Toxin Producing Escherichia coli). E. coli O157:H7 has two stx toxins, stx1 and stx2.

8. How does it cause disease? E. coli O157:H7 is an EHEC strain which kills epithelial cells in the gut, resulting in bloody diarrhea. It also invades the urinary tract causing an ascending infection which damages the kidneys. But it gets worse. Broad spectrum fluoroquinolone antibiotics such as ciprofloxacin which are often used to treat infections cause an SOS response in E. coli cells which in turn induces the lytic cycle of the lysogenic toxin-carrying phages. This results in a thousand-fold increase in toxin expression. Treatment with some some beta-lactam antibiotics also increase stx toxin production.

9. Many people recover without antibiotics or other specific treatment in 5–10 days. There is no clinical evidence that antibiotics improve the course of disease, and some may make it much worse (see above). Haemolytic-uremic syndrome is a life-threatening condition usually treated in an intensive care unit. Blood transfusions and kidney dialysis are often required. Even with intensive care, the death rate for haemolytic uremic syndrome is 3%–5%.

10. Wash your hands thoroughly with soap and warm water after contact with animals. Wash raw vegetables such as salads well before eating. Cook meat thoroughly all the way through, especially burgers and sasuages where external contamination of meat is transferred to the inside by mincing.

11. E. coli is a Gram-negative bacterium. IT’S NOT A VIRUS! So next time a journalist talks or writes about “the E. coli virus” – do us all a favour and yell at them!

Related:

• The continuing evolution of E. coli O157:H7
• E. coli O157:H7 – getting to the bottom of the burger bug
• Bacterial toxins

The fungal kingdom is vast, spanning 1.5 to as many as 5 million species diverse as unicellular yeasts, filamentous fungi, mushrooms, lichens, and both plant and animal pathogens. The fungi are closely aligned with animals in one of the six to eight supergroups of eukaryotes, the opisthokonts. The animal and fungal kingdoms last shared a common ancestor 1 billion years ago, more recently than other groups of eukaryotes. As a consequence of their close evolutionary history and shared cellular machinery with metazoans, fungi are exceptional models for mammalian biology, but prove more difficult to treat in infected animals. The last common ancestor to the fungal/metazoan lineages is thought to have been unicellular, aquatic, and motile with a posterior flagellum, and certain extant species closely resemble this hypothesized ancestor. Species within the fungal kingdom were traditionally assigned to four phyla, including the basal fungi (Chytridiomycota, Zygomycota) and the more recently derived monophyletic lineage, the dikarya (Ascomycota, Basidiomycota). The fungal tree of life project has revealed that the basal lineages are polyphyletic, and thus there are as many as eight to ten fungal phyla. Fungi that infect vertebrates are found in all of the major lineages, and virulence arose multiple times independently. A sobering recent development involves the species Batrachochytrium dendrobatidis from the basal fungal phylum, the Chytridiomycota, which has emerged to cause global amphibian declines and extinctions. Genomics is revolutionizing our view of the fungal kingdom, and genome sequences for zygomycete pathogens (Rhizopus, Mucor), skin-associated fungi (dermatophytes, Malassezia), and the Candida pathogenic species clade promise to provide insights into the origins of virulence. Here we survey the diversity of fungal pathogens and illustrate key principles revealed by genomics involving sexual reproduction and sex determination, loss of conserved pathways in derived fungal lineages that are retained in basal fungi, and shared and divergent virulence strategies of successful human pathogens, including dimorphic and trimorphic transitions in form. The overarching conclusion is that fungal pathogens of animals have arisen repeatedly and independently throughout the fungal tree of life, and while they share general properties, there are also unique features to the virulence strategies of each successful microbial pathogen.

Microbial pathogens in the fungal kingdom. (2011) Fungal Biology Reviews 25(1): 48-60

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