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Monday, 12 February 2018 19:40

TWiV 480 Letters

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Published in Letters

Trudy writes:

I have these on my Amazon wish list and plan to buy them sometime soon, so I haven’t read them yet! However, I found out about them because one of the authors was on Meet the Microbiologist.

https://www.amazon.com/gp/product/0545044227/ref=ox_sc_act_title_3?smid=ATVPDKIKX0DER&psc=1

https://www.amazon.com/gp/product/0545273226/ref=ox_sc_act_title_2?smid=ATVPDKIKX0DER&psc=1

https://www.amazon.com/gp/product/0545805414/ref=ox_sc_act_title_1?smid=ATVPDKIKX0DER&psc=1

Steve writes:

Hi Vincent et al,

Just listening to this week’s TWiV, and should have been asleep hours ago, but had to pass this link on to you. As I’m in the UK, my search yields UK links, but I should think you can get this ‘colouring book’ in the US too.

It looks like it is just a child’s book, but the botany is all good, and the diagrams are useful for anyone beginning botany, or just loving plants. I’ve had it for years, and tended to photocopy the pictures to share round for people–not just children–to colour in. Colouring is also surprisingly popular with adults nowadays, so there are a number of good books like this one around.

All the best (And you certainly do deserve that award that was mentioned earlier in the program–I’ll be looking at that later…),

Steve

Luton

Bedfordshire

England

https://www.worldofbooks.com/botany-coloring-book-by-paul-young-gor001550982.html?keyword=&gclid=CjwKCAiA-9rTBRBNEiwAt0Znw6vJzcoQbjPkIRJs1SHlqB_TsgeORZPyVTE4EBnc2aIC3F7m2rfxPhoC2E4QAvD_BwE

Bjørn writes:

Dear twivites, all of whom I love and admire from a distance

In this recent PNAS paper about flu “Infectious virus in exhaled breath of symptomatic seasonal influenza cases from a college community”

     http://www.pnas.org/content/early/2018/01/17/1716561115

the following is stated: “Fine-aerosol viral RNA was also positively associated with having influenza vaccination for both the current and prior season.”

That’s an interesting statement for many reasons. It sounds controversial, but it could also  be true even if not very controversial. However, I couldn’t tease that detail out of the paper.  Here are a couple of interpretations, could you help me determine which one is correct?

  * “In a population of infected individuals, individuals that recently got a vaccine are more likely to shed  aerosolised virus”.

  * “In a general population, individuals that recently got a vaccine are more likely to shed  aerosolised virus”.

The exact answer to both of these questions will depend on the efficacy of the vaccine, and the actual difference in amount of shed virus between those who got it and those who didn’t. Maybe the answers to both are in the paper, directly or indirectly,  but I  couldn’t  find them.

However, I’d like to know for sure because if  the second interpretation, for whatever reason, is adopted by the antivaxers, the consequences will be dire since it could be interpreted as “peer reviewed evidence that flu vaccines are harmful”, which could possibly make more people refrain from having vaccines that would help both themselves and those around them. A recipe for neutralizing antibodies, in the form of fact based interpretation of the paper’s data,  to stop this meme from “going viral” would be useful. Can you help?

In addition the paper says a number of interesting things about the nature of aerosolised flu virus in exhaled breath, but it’s also a technical paper so it’s hard to parse for a computer scientist type like myself without any other background in microbiology than a few online courses, including Vincent’s excellent online courses on viruses, and a steady diet of TWIx podcasts

You are, as always, an inspiration.

Best Wishes

Bjørn in Oslo, Norway

Christoph writes:

Dear TWIVsters,

Thank you for the comprehensive overview of our bioRxiv manuscript on the Bodo saltans virus. Dickson represented the Protistology part of the manuscript well and it was great to hear his excitement about the topic! bioRxiv has been an interesting experience and we have received comments from a number of individuals from different backgrounds on the preprint. The in-depth discussion on TWIV was helpful to identify sections in the manuscript that need clarification for the non-specialist audience. We especially would like to thank Kathy for sending us her written comments. Overall, the TWIV comments nicely complemented the peer-review feedback and will allow us to ultimately write the best possible paper.

That being said, we would like to address a few points raised in the episode:

  1. The isolation process and how we were able to find a virus for Bodo saltans: To isolate viruses, we sampled 11 freshwater environments (and many more marine ones) and isolated hundreds of potential host organisms from these environments. The pond on campus that yielded Bodo saltans was one them. We pooled and concentrated the viral fraction from all samples and screened it against the potential hosts. Therefore, we are certain about the origin of the host, but can only say that the virus was in one of the 11 samples. We were not able to reisolate or detect the original virus in the pond, but my guess would nevertheless be that the virus came from the same sample that the host is from.
  2. The carbohydrate metabolism manipulating genes: It was great how excited Dickson got about this! Similar genes to the ones in our virus are actually found in many giant viruses. They have been best described in Chloroviruses, check this wonderful review by James Van Etten et al. entitled “Chloroviruses Have a Sweet Tooth”: http://www.mdpi.com/1999-4915/9/4/88/htm
  3. The mobile elements in essential genes: This is complicated, but allow me another attempt at explaining this. There are three levels of activity to consider: The inteins, as autocatalytic proteases, can excise themselves from the precursor-protein. The group-I introns, as autocatalytic ribozymes, can excise themselves from the pre-mRNA. On top of all this activity, and this is I think where the confusion stems from, inteins and introns possess internal open reading frames that encode homing endonucleases (red in Fig 4). These endonucleases are sequence-specific and induce double-strand breaks in the target location, which then leads to the insertion of the mobile element in a new locus. This is what makes these introns and inteins mobile genetic elements by allowing them to spread through the genome. The target site of the endonucleases is usually in essential genes of a single copy, such as the DNA polymerase gene, which allows efficient invasion. Due to the excision of the element, either on mRNA or protein level, the gene product is still functional. Any imprecise excision or modification of these elements would disrupt the coding sequence of the essential gene and thus be “lethal” to the virus. Since only a clean and precise deletion of the entire element would restore the original gene, the host is pretty much incapable of removing the this genomic parasite and is thus forced to carry it along. A great review on the topic by Gogarten and Hilario can be found here: https://bmcevolbiol.biomedcentral.com/articles/10.1186/1471-2148-6-94
  4. The environmental representation plot: The way this program (pplacer) works is that it attributes the environmental metagenomic sequences, in this case from the TARA oceans survey (http://ocean-microbiome.embl.de/companion.html) to the most similar sequences in the reference tree. So effectively, we are not saying that our specific virus is super abundant in the ocean, but rather that most of the giant viruses in the ocean are similar to BsV, CroV and the Klosneuviruses.

These and all other issues will be addressed in the final paper, so to borrow from TWIM, “stay tuned”.

Thanks for all the great work you do on the TWIX empire!

Best

Chris and Curtis

Ben writes:

Dear TWIV hosts,

I was recently listening to episode 474 in which you were discussing viral infections in the ocean and how viruses kill ~20% of marine microbial standing stocks every day. Dickson stated that the tropical oceans were ‘clear as a bell’, and thus largely sterile (what Dickson meant to say was “nutrient poor”. Sorry for the misstatement). In fact the open oceans are far from sterile – surface water in even the most nutrient-limited parts of the subtropics contain ~1 million bacteria per mL of seawater. These communities are dominated by two bacterial groups – Prochlorococcales, which are phototrophic, and the Pelagibacterales, which are heterotrophic. While the Prochlorococcales are restricted to the photic zones, the Pelagibacterales are found at all depths in every part of the global ocean, and comprise up to 50% of all marine microbial communities. Such is their abundance globally, that their biomass is estimated to be similar to that of all marine fish. They are in all likelihood the most abundant bacteria on the planet. As heterotrophs that specialise on metabolism of small, labile carbon compounds, including those released from the high degree of viral lysis discussed in the episode, it is estimated that a large component of total global primary production is converted back to atmospheric CO2 by the Pelagibacterales (see here for a great review: http://www.annualreviews.org/doi/10.1146/annurev-marine-010814-015934).

Apart from their abundance, these organisms are biologically fascinating for reasons touched on in the same episode – that of genome size and ‘overhead’. Members of the Pelagibacterales possess some of the smallest genomes of any free-living organisms (~1.3Mb and encoding ~1350 genes). They are small enough to pass through a 0.2µm filter, with a cellular volume ~ 1/60th of an E. coli cell (their genome takes up ~30% of their cell volume). To minimise costs of genome replication in nutrient-limited waters, they encode almost no regulatory proteins. Consequently, almost all their proteins are either constitutively expressed, or regulated by riboswitches. In some cases, metabolic pathways are regulated by kinetics, switching from one to the other as intracellular concentrations of metabolites change (https://www.nature.com/articles/nmicrobiol201665/fig_tab). Evolution in nutrient-limited oceans has also resulted in an AT-rich genome (~27% GC), which reduces the nitrogen requirements for DNA replication.

While they are challenging to culture, we can now do so thanks to decades of efforts of Prof. Steve Giovannoni (Oregon State University). They don’t grow on plates, and need to be grown in either polycarbonate or teflon plasticware in liquid culture. They are auxotrophic for numerous compounds including reduced sulfur and thiamine precursors (again, reducing the cost of genome replication has made them highly dependent on the outputs from their community neighbours due to loss of function). They can’t be grown on rich medium, possibly due to a lack of regulation to control the flow of unbalanced metabolites to central carbon metabolism – they grow best when, for instance, glycine and pyruvate are provided in specific stoichiometries (https://www.nature.com/articles/ismej2012122). Their slow, constitutive lifestyle means that they tend to get outcompeted by ‘weeds’ like Vibrio and Pseudomonas. So, the only way to isolate them is to put a single cell in a few mLs of medium and let them grow with no competitors. This can be an exercise in patience because they double every 48 hours or so. With the right know-how, we can now culture them to ~10^8 cells per mL. Even at these densities (measured with flow cytometry), to the naked eye the cultures look no different to sterile control medium as the cells are too small to impact much on optical density (they are also kept in suspension by Brownian motion, so no need for shaking). This perhaps explains why tropical seawater is so clear, even though it’s full of microbes.

Coincidentally, it was my fascination with the biology of these organisms and the evolution of ‘streamlined genomes’ that led me through a circuitous route to TWIV. I was a postdoc in the lab of Steve Giovannoni when another postdoc there (Yanlin Zhao) isolated 4 new phages that infected the Pelagibacterales in 2013. As a bioinformatician, I was tasked with seeing how abundant these new phages appeared to be in global ocean metagenomes. After some fairly rigorous analyses to make sure we weren’t mistaken, it turned out that these viruses were probably the most abundant viruses on Earth (https://www.nature.com/articles/nature11921). Up until that point, the high abundance of Pelagibacterales had been thought by some to be due to ‘defense specialism’, where its size and slow growth allowed it to escape viral predation. The work showed that even in the age of metagenomics, culturing is still important. Microdiversity in these phages limits assembly from metagenomes and their genomes tend to fragment into small pieces that are often discarded (https://www.nature.com/articles/ncomms15892). Once we knew what we were looking for using genomes from cultured viruses, we could find their genomic signatures everywhere we looked. During this project, and somewhat naive to the world of virology, I devoured every available resource I could find, including TWIV, which I have been listening to ever since. Once the door of virology was open, I’ve never looked back and directed my own research to better understand host-virus interactions and how they shape microbial communities.

I am now a new PI at the University of Exeter, UK, where I teach bioinformatics and marine microbiology and virology. My small group is currently developing new sequencing methods for long-read viromics using the new Nanopore sequencer, with the capacity to capture near-full-length dsDNA viral genomes on single reads. This new approach also overcomes the microdiversity assembly issue. My awesome PhD student who did most of this work, Joanna Warwick-Dugdale, is currently writing up the manuscript for biorxiv. I have recently got my first major grant (NERC-funded for 4 years) to investigate host-virus interactions in the Pelagibacterales to provide some much-needed insight into how these viruses shape Pelagibacterales populations and evolution, as well as their impact on global carbon biogeochemistry (and will be recruiting postdocs for this project shortly!). One hypothesis we will test is that specialisation on small carbon moieties that result from viral lysis allows the Pelagibacterales to dominate communities, even in the face of extreme viral pressure, because they are highly efficient at scavenging from their own dead.

Thank you for all you do to promote science to such a broad community with your podcasts! They’ve taught me a great deal about virology outside of the marine environment and are a staple part of my morning commute.

Best wishes,

Ben

Steve writes:

Hi Vincent et al,

Just a quick note after listening to your brief sortie into the gender minefield. ‘What is a ‘girl’? ‘

You might be surprised to hear what Stephen Fry and the QI team had to say on the subject:‎

https://m.youtube.com/watch?v=h9iYvSztyHs

Always impressed and entertained,

Steve

Luton

Beds

UK

James writes:

Hi TWiV team!

I just listened to your special episode with Ray Ortega about being a podcast producer. I am very interested in this still evolving form of communication, so although there was little to no actual science or virology discussed, it was one of my favorite episodes so far because I got to listen to a behind-the-scenes take in what goes on in the podcasting world.

I am writing in response to comments on the show about podcast length. I may be (probably am) in the minority here, but I listen to many podcasts and generally prefer shorter ones—15 to 40 minutes—because it fits with my lifestyle better and allows me to get an overview of the topic that I can take a deep dive into later if I choose. I personally tend to listen mostly when I am commuting or performing mundane and repetitive tasks in the lab like streaking or pipetting. So I don’t really “settle in” so much as look for a way to learn on the go and expand my knowledge in a variety of topics.

I’m not saying you should change anything. I’ll listen even if the shows become longer! Just wanted to provide my thoughts so that all views may be represented.

One other thing: I like episodes where there is more focus on news and current events in virology, microbiology, and science. For example, the recent episode where the team discussed the ban on gain-of-function studies, and the opening of a new BSL-4 lab. I especially liked how these were discussed first, before the papers. This gives me a great chance to hear from the experts and share these accessible topics with friends and family.

Thanks for the great shows and I look forward to each and every one!

Vincent Racaniello

Vincent Racaniello, Ph.D. is Professor of Microbiology at Columbia University Medical Center. As principal investigator of his laboratory, he oversees the research that is carried out by Ph.D. students and postdoctoral fellows. He also teaches virology to graduate students, as well as medical, dental, and nursing students.

Vincent entered the world of social media in 2004 with virology blog, followed by This Week in Virology. Videocasts of lectures from his undergraduate virology course are on iTunes University and virology blog. You can find him on WikipediaTwitter, Facebook, and Instagram. His goal is to be Earth’s virology professor. In recognition of his contribution to microbiology education, he was awarded the Peter Wildy Prize for Microbiology Education by the Society for General Microbiology. His Wildy Lecture provides an overview of how he uses social media for science communication.

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