Live-blogging from the Nobel Dialogue conference was a new experience for me—the aim being to sit through a lecture/dialogue, digest the information, write a coherent and constructive blog post and then publish before (or while) heading to next discussion. Despite some thorough research prep ahead of the first day, and reading plenty of live-blogging advice about battery power and staying hydrated, I managed to arrive in the auditorium totally dehydrated, and proceed to deplete my macbook battery in 90 mins. Thus, my first post was executed by mobile phone on WordPress for mobile and submitted by sacrificing my attention to the next talk. Fortunately, the nice thing about having a team other other bloggers is that you know they have the next talk covered.

The real struggle was doing likewise in the panel discussions. The panels consisted of five/six eminent thinkers, expertly moderated by Matt Ridley (for Genetics in Agriculture and the Environment) and Adam Smith (for The Promised Land of Genomic Medicine); however, despite excellent moderation, you’re still left with the task of weaving multiple streams of consciousness into a cognate narrative. After the first panel there were roughly five minutes to consolidate four pages of notes into a post and be ready to engage with the next discussion. I just about managed that, but the final panel session still awaits my finished post because, by that time, I was as fried as my macbook.

Covering the Nobel Dialogue was, however, a great experience. The organisation and slickness of this conference was superb, and audience participation and access to panelists (especially during coffee/lunch/wine breaks) was unfettered. I’ll write more about my thoughts on the Nobel arena in the coming days, while I finish up posts and overall synopsis. For now I’m cross-posting the first lecture of the day I covered (written on my mobile).

Helga Nowotny on the promise of science

Nowotny asks what does a social scientist do in the land of genomic promise? The promise (or contract) that binds science and scientists together is the expectation that the public have from scientists, and the scientists aspiration to produce something useful. The pursuit was one of laws, the laws of natural philosophy, but with the concept of ‘law’ as a term taken from society. Yet the aim was to supersede the laws of society; science promised access to the kind of laws that could not be bent by kings or rulers – they provided truths above the arbitrariness of the rules of society. For the early scientists, Francis Bacon et al. the promise was the affecting of all things possible, using the experimental method and practical objectives, for the betterment of humankind.

Early promises that have been realised in society include the reduction in working hours with the concomitant provision of more free time, which ironically we fill with the products of other scientific promises, those of technology – mobile phones, TV, travel. As we consider the impact of genomic science, the genomic promise is a global promise where investment flows globally and through decentralised networks. Is genomic science now in the position to take the promise of human enhancement/betterment to a new level?

Nowotny quotes Richard Powers –  “People want everything, that’s their problem”

With genomics we may be entering a second renaissance. Like the original renaissance that put the individual at the centre of the political and cultural world, genomics offers the opportunity to recognise and discover our own individual uniqueness in a deep sense together with our relatedness to each other and to other species.

However, we need to be careful with promises we make, and not betray them. The worry is that we arrive in a situation where post-genomic science ends up meeting a ‘pre-genetic society’ that isn’t ready for it. There need to be bridges that enable society to be ready for these new technologies; these, Nowotny explains, could include citizen science projects. Create space for people’s everyday experience and integrate it as experiential evidence. One such project is personal genomics – allowing people to interface with and explore the question, ‘who am I?’

From my own perspective, and reflecting on Nowotny’s ideas of promise and ‘betrayal’, it’s worth reflecting on that fact that the human genome project was funded off the back of such a promise – that benefits would be delivered upon its completion.

That we needed the genome sequence goes without saying, but the fact that most scientists knew the answers would not (could not) be immediate was probably known to scientists in a way that it was not clear (or not made clear) to society, or the politicians ratifying the funding. Although we could ponder on whether a government would have provided the funding it did had it known it wouldn’t get immediate results.

If the promise of science in society is one of expectation, I think it’s probably important to be careful managing those expectations. These begin with education about how science works, it’s pace and the absolute need to establish the fundamentals before we seek to apply discoveries to society.

This post originates from the Nobel Week Dialogue 2012, where the author is a member of the official blog team. Follow the official event blog here.

[This post was restored from a WayBackWhen archive. It was originally posted to a blog called ‘The Gene Gym” that began life on the Nature Network in 2010, and then moved to Spekrum’s SciLogs platform. Unfortunately many of the original images have not survived the import]

It’s amazing how far you can travel internationally before smelling fresh non-air conditioned air. I arrived at Arlanda airport – 50 km North of Stockholm – in the late afternoon, and was immediately siphoned into the familiar human corral of border security that is facsimiled the world over. The fastest route from the airport to the city centre is via the Arlanda Express, a name that evokes a surety of function that I’m willing to accept as a factual statement in this very modern Scandinavian country. Sure enough, in a mere 20 minutes I am delivered to Stockholm central station and can make my bid for the open air; yet emerging into a fog of -9C air was probably rather more fresh than I’d anticipated. Stockholm is seasonably festooned with lights, which twinkle in the crystalline cold and cast their light on streets paved with a compaction of snow, sand and salt, giving the consistency of gingerbread dough.

I’ve arrived in a city that must be full of Nobel Laureates right now, enjoying some of the pomp and ceremony of Nobel week festivities ahead of the prize givings on Monday 10th December, and indeed some of whom will be at the Nobel Week Dialogue tomorrow. This is after all why I’m here, and yet what the dialogue will bring I’m not sure. There is certainly the potential for great content, but it will depend very much on the moderation. It’s going to to be an intriguing one to blog about, not least because the format and frequency of blog has not been established, because we don’t know quite how the panel sessions will run. With any luck key themes-within-a-theme will emerge and provide a narrative for content.

I’m only here for a short time, so I’m going to make every effort to capture something of the delights on offer. I’ve cross-posted an interview I had with Kerstin Lindblad-Toh, a very talented scientist whose research output and current roles are prodigious. I hope to cover more of her work in the future. There will be more cross-posting in the coming days, but in the meanwhile I leave you with a first image of Norra Bantorget square, Stockholm.

Credit: © Jim Caryl, 2012

An important means by which we try to understand the human genome is, oddly enough, by looking at the genomes of other mammals. The aim is to identify areas of evolutionary constraint, regions of the genome that we all share (both coding and non-coding) and are thus likely to be important for all of these species. These regions have not only assisted with the identification and assignment of genes on the human genome, but they also provide important information about disease associated mutation. One mammal genome, unique amongst the others, offers particular insight into our genes and inherited genetic disease, the domestic dog.

The dog genome, published in 2005, was the fourth mammalian genome to be completed (after man, mouse and rat); the dog occupies a curious position in nature as it has shared a mutually beneficial relationship with humans for at least 15,000 years, living in the same environments as us and sharing our food. Over this time humans have selectively bred dogs for companionship, hunting, shepherding and other uses, and in so doing have channeled the diverse canine genome into a variety of behaviours, shapes and sizes.

This selective breeding has resulted in dogs having a ‘simplified’ genetic architecture, but has inevitably made them a storehouse of some 400 inherited diseases, many of which are shared by humans.

I contacted Nobel Week Dialogue participant Professor Kerstin Lindblad-Toh, whose contributions towards mammalian genome projects span the mouse, dog, horse and short-tailed opossum. I spoke to her about her ongoing work based on the dog genome, to find out more about the contribution of ‘man’s best friend’ to science and society, and the other work with which she’s involved.

Q. The domestic dog genome work seems to have benefitted from of the particular importance of dogs in society, and given that they are prone to a number of shared diseases it seems appropriate that this relationship can also benefit human health. I appreciate that all this work is the product of collaborations, but I’m curious about how the dog genome studies actually got started? Was there a particular demand from groups/societies focussed on dogs? 

Kerstin: The early dog research was advocated by Jasper Rine [UC Berkeley] and Don Patterson [Rtd. University of Pennsylvania] and funding by organisations such as the American Kennel Club/Canine Health Foundation. In 2002, I was contacted by a group of researchers, led by Elaine Ostrander, interested in dog genetics and wanted the genome sequenced. We (I) then developed a plan to sequence the genome where we also surveyed the variation in the genome. This work was funded by NIH for its comparative value and was performed under my leadership at the Broad Institute.

Q. How do we actually translate disease information from these studies? Are there key findings from your comparative genomics work that are being successfully translated into human patient studies?

Kerstin: Examining the human genome and understanding the function of every gene and how they are turned on and off correctly is challenging. We try to address this in two different ways: First we compare the genomes of many mammals to find elements that are similar across all species. These must be important/functional as nature has conserved them. Secondly, we know that finding disease genes is easier in dogs than humans. Therefore we find novel disease genes in dogs and then we examine the same genes in human patients. We have very exciting results where genes found in lupus, autoimmune inflammatory diseases and lymphoma respectively is teaching us about the corresponding human diseases.

In addition we find genes that define specific pathways involved in obsessive compulsive disorder.

Q. So clearly the sequencing of the canine genome will have a significant role to play in understanding the basis of shared diseases between dogs and human, but I also see that a number of your papers reflect on an area of more fundamental pursuit, that of the evolutionary history of dogs. Is an area of academic writing that has a particular interest?

Kerstin: [these areas] can reveal exciting information about canine evolution, and perhaps its connection to human evolution, we have sequenced the genomes of many dogs and wolves and compared them. We find signatures of selection that show which types of genes have been important for dogs to adapt to living with humans.

Q. It’s tempting to speculate that by selectively breeding an animal to have familiar human-interactive traits—such as a dog’s ability to recognise social stimuli—we can look at those regions that are changed compared with ancestral lineages (such as the wolf). Taking this further, can we derive from this a tantalising glimpse of those parts of our genome that reflect those same traits in us?

Kerstin: I believe that there will we regions of the genome associated with behaviour and the ability to read human social signals. In addition to comparing dogs and wolves to detect genomic regions of importance. We are also comparing working dogs that have been carefully tested for behavioural traits. We hope to find genes underlying positive behaviours such as curiosity and sociability but also for more pathological conditions such obsessive compulsive disorder. I believe that many of the genes and mechanisms underlying behaviour will be shared between man and his best friend.

Q. An area of genomics that is increasingly being featured in science reporting is that of the roles of epigenetic DNA modification and 3D chromosomal architecture in regulation and function of genomes. To what extent are you also comparing animal and human ‘epigenomes’, is this a feature of current studies?

Kerstin: Epigenetic signal of course have great importance for genome regulation. While others in the field have started on these studies, we have not done so ourselves yet. While epigenetics should also be studied, I believe it is the inherited mutations and the breed structure that makes dogs unique.

Q. At the Broad Institute you are responsible for the Mammalian Genome Project, which includes the 29 mammal project. How has the project chosen the 29 mammals? Why these particular mammals?

Kerstin: The 29 mammals project brings important information about the genes and regulatory elements in the human genome. The first few mammals (including mouse and dog) were chosen for their biomedical relevance. The next 24 mammals were chosen primarily to provide a broad sampling of mammals to bring as much evolutionary divergence, contributing power to the project. When two mammals contribute equal power a species with biomedical relevance would of course be chosen.

Q. The early genome sequencing projects heralded an age of ‘big data’, and in the past our ability to analyse such volumes of data has struggled to keep pace. Are the tools you now have available for looking at big data finally starting to keeping pace? Or has the process by which research is undertaken in these fields had to change?

Kerstin: The amount and complexity of data generated is amazing and is continuously growing. At SciLifeLab we have the competence and resources to design, generate and analyse innovative experiments. We strive to make use of unique human or canine patient materials and to ask questions of relevance to translational medicine. Due to the continuously changing arena both in technology and analytical methods we need to be both nimble and innovative.

Q. In terms of careers, I’m curious about your route to where you are now? Early career scientists are often interested to hear how established scientists have arrived where they are. 

Kerstin: After a basic education in molecular biology at Stockholm university I received my PhD in human genetics at the Karolinska Institute. My postdoc time was spent with Tom Hudson at WI/MIT, where I had perfect combination of freedom and a supportive advisor when needed. I performed multiple genomics projects involving both disease mapping and technology development. One day I ventured into Eric [Lander]’s office with an idea, and apparently my timing was right, because he asked me to lead the mouse genome project. This project emphasised the value of interdisciplinary collaborative projects and made me realise the need for a better understanding of the human genome and the value of comparative genomics, making me determined to realise these goals.

Q. You currently have appointments both at the Broad Institute at MIT, Uppsala Universitet and SciLifeLab. Senior appointments in any one place would take a huge amount of time and effort, not least to keep on top of research projects, and writing papers/grants. How do you manage? 

Kerstin: This is challenging, but with good colleagues, hard work and efficient planning it is possible. I spend roughly half a year in each place, with some travel back and forth. Either way I spend a lot of time on the phone.

Q. Finally, the nwd12 conference will bring together many faces familiar to you, as well as potentially new ones. Are there any particular messages – for your own part – that you’re aiming to communicate, and are there any discussions you’re particularly looking forward to?

Kerstin: I’m really hoping to convey the message of how important the non-coding regions of the human genome are. The regulatory portions of the genome facilitate evolution and adaptation to novel environments as well as underlie the vast majority of our common diseases.

This post originates from the Nobel Week Dialogue 2012, where the author is a member of the official blog team. Follow the official event blog here.

On Saturday morning I’ll be rising at a hideously early hour to board the first of two flights that will (eventually) bear me on my merry way to Stockholm. I was lucky enough to be invited to be part of the blog team covering the Nobel Week Dialogue 2012, a new annual conference that will form part of the week of events leading up to the Nobel Prizes. I’m a big fan of conferences, I’ve lost count of the number I’ve been to over the years, but often classify them into two camps:

1. Specialist conferences: where I’ve gone to strut my academic stuff and watch others within my research community strut theirs – and when off the dance floor, we also listen to each other talk; and,

2. Geek/fan conferences, where you go immerse yourself with kindred spirits who share your interests, or where throw yourself into something new to learn and experience.

It’s a subtle distinction, I know, given the large number of fellow ‘specialists’ I also encounter at fan conferences, but the less said about those Star Wars/Trek conventions the better. I’m going to put the Nobel Week Dialogue in my fan conference partition, because I think I will be amongst kindred spirits who are all eager to explore the what on earth the genomic revolution means in today’s society.

‘Big Data’ has been the buzz word for some years now, partly because in the past five years we’ve generated more data that the entirety of human history, but how can we realise the promise of such technologies to advance medicine, agriculture and the environment? It’ll take a concomitant adaptation of all the other institutions of society too, and I don’t know what’s trickier? Science has had a head start in the game, but policy makers, ethicists and business leaders are going to have to get caught up, lest the promise and investment become a purely academic exercise.

I’m certainly not attending as any kind of ‘expert’, such as there can be any ‘experts’ in this BIG subject, but this conference is a dialogue between leaders of several ‘big data’ fields, scientists, policy makers, thinkers and interested members of society, and I count myself amongst several of those. I want to know, I’m genuinely curious, and I wonder who really has a grip on it all? When I know, you will, because I’ll be writing about it. I’ll also be cross-posting some of my posts over the weekend, but also take some time to check out the work of the other bloggers covering the conference on the conference blog.

The conference wil be live-streamed too, and whilst you’re waiting, look at the participants – the meta-CV of all these heads is mind-boggling!

I’ve copied my first, introductory, post below. Next stop, Stockholm.

I have been an active research scientist in bacterial genetics for over a decade, and a science blogger for about four years, with much of my research and writing focusing on various aspects of the lifestyles (and DNA) of bacteria. Yet in all the many hours I’ve devoted to staring at Gs, As, Ts and Cs of DNA on a screen, I’ve never personally made use of the information to understand my own genome – something I’ve been increasingly interested to investigate.

It seems an odd admission; I’ve been an early adopter of every other nascent technology – both for personal and research use – but I guess the major reason for not investigating my own genome is the question: can we really understand it yet? So I’m quite excited to have an opportunity to visit Stockholm (for the first time) and attend the Nobel Week Dialogue. The 2012 Nobel prizes marks 50 years since the 1962 prize was awarded to James Watson, Francis Crick and Maurice Wilkins for their ‘discoveries concerning the molecular structure of nucleic acids and its significance for information transfer in living material’; or as they once put it – (discovering) ‘the secret of life’. It is on this theme that the Nobel Week Dialogue will focus, bringing together a broad range of scientists, thinkers and policy makers to examine the genetic revolution and its impact on society.

I feel extremely fortunate to have worked in molecular sciences over the past decade. The first working draft of the human genome was completed during my first year in graduate school. The race between the private company Celera and the publicly funded HGP to sequence the 3 billion letters of our DNA had been very much the competitor sport to watch during the course of that spring. Each subsequent year seemed to bring a whirlwind of technological advances and new understandings in DNA technology, improving the speed, and lowering the costs of DNA sequencing. By the time I left my most recent research lab we were able to sequence entire bacterial genomes (all be it squished together on a single sequencing chip) in just over a week for £50 a piece – an unthinkable speed and price only a few years previously.

So the question remains, is this the right time to have a look at my own DNA? Admittedly the personalized genomic industry isn’t sequencing your entire genome; rather they give you an overview of genes with known ‘phenotypes’ – those characteristics that can be observed. Some of these may just reveal curiosities – such as why you may be particularly affected by caffeine – or may reveal more disturbing findings, such as a pre-disposition to a certain disease; I can’t deny that I have a nagging curiosity about the former, but have less inclination to discover the latter. I’m also aware that just knowing a DNA sequence doesn’t necessarily tell you whether, how and when it is used; all cells ultimately contain the same code, and yet a nerve cell is able to extract from the code an exact set of instructions for how it should look and function, and likewise a liver cell. How two such different cells can read from the same material, yet grow and differentiate into such different roles – and remain that way – is key to understanding both how organisms develop, but also to how errors in these processes can result in diseases such as some forms of cancer.

I’m hoping there will be some great discussions on how our growing understanding of these processes can be applied to ‘genomic medicine’, and as I will be live-blogging the afternoon session 1B (The promised land of genomic medicine: how do we actually get there?) this will likely focus on the socioeconomic and ethical considerations of what we learn, and how we get there. I’m also live-blogging the afternoon session 2A (Genetics in agriculture and the environment: where can the science take us?), and I’ll be interested to see which of several different directions this takes, but with Matt Ridley as moderator, I’m sure it’ll be engaging.

I am grateful to the organizers for the opportunity to attend and write about the Nobel Week Dialogue, and I can’t deny that I hope to get to see something of the city – thinly veiled in the winter light as it will be in December.

This post originates from the Nobel Week Dialogue 2012, where the author is a member of the official blog team.

When I was an undergraduate you couldn’t get a journal article online, about the most you could hope for was a table of contents (ToC). Getting access to scientific articles meant a visit to the library with a photocopy card and a great deal of patience. ‘Bagging’ a photocopier was an artform in itself – I think everyone thought they knew of a secret, hidden photocopier on an upper level of the library, one that you felt you owned. Of course, this fog of solipsism would evaporate upon arriving to find your photocopier being used by another person. You would then resign yourself to waiting, staring indignantly at the interloper.

There were those students who were either too lazy, or too apathetic to perfect the art of photocopying; these people were destined to long, long waits and having to find new and inventive ways of marking their presence in the queue while they used the bathroom. Of course, you could make a few pounds (or free pints) by selling your pristinely photocopied articles to them. I think everyone experienced those occasions where, within reach of the photocopier, you would find that the article you thought you had was actually in the next bound edition along on the shelf, or you’d finally arrive to find the toner so depleted that you would need to pencil over some of the words in  to make them legible. That was if you were lucky – if you were unlucky, you could expect there to be a paper jam, or no paper left.

Access to articles was also limited to which journals your university subscribed to in print. During my honours research project, if any of the PhD students/postdocs in that lab heard that someone was planning a library visit to Manchester or Leeds, they would not escape without a long list of journal article requests from them, and a £10 note by way of compensation for a weekend of pain in an unfamiliar library, ignorant of the pecking order.

So why am I telling you about this? Well, I have long considered myself an advocate of open access (OA) publishing, and in fact my lab went to some considerable expense to pay for my first publications to be OA, several £thousand in fact. I welcomed all efforts towards OA journals and celebrated the day when the PLoS journals came into being. In a frank admission, however much I supported this cause I’d never actually experienced being without access. For 16 years, from the first day I set foot within a university, I have always had access to journals. As I worked in a large Russell Group university for the entirety of my research career, I have never wanted for an online article; I was often unaware when articles were behind a paywall because my computers both at work and at home were set up to pass them without hinder.

I recently finished my latest research contract, and within a fraction of a second of leaving the department I lost all access to my former employer’s eResources. For the first time in my life I’m unable to access 80% of the articles I need without having to email friends and request them. As I suddenly find myself trying to read up and prepare to be part of the blogging team at the Nobel Week Dialogue in Stockholm in December (more about that in an upcoming blog post), as well as do some contract consultancy and prepare for a pretty major interview, I’m being blocked at every turn. It makes me wonder how the hell professional freelancers do it? For amateur enthusiasts this is an annoyance, making them reliant on some of the great science writing available online, but to jobbing scientists and freelance science writers trying to keep up with their game, it’s like having a door slammed in your face.

I can only think that, in principle, the UK Gov Plc getting behind OA publishing is a good thing; however, I have every expectation that they’re merely going to pay for it by taking money away from research funding, rather than providing an extra quota for it. I find it all the more annoying that if I didn’t already have copies of my papers, I would not be able to access some of those I wrote! Perhaps, at the very least, journals for whom we peer review could give us permanent access? I was interested to see that in the latest paper I reviewed, the review process itself gave me temporary access to a whole range of journals, a fact that I’d not noticed before.

I’ve found a temporary solution, and remain hopeful that – all things going well – I should have access restored, however, for a great many professionals and amateurs alike the presence of paywalls for accessing material that WE scientists produce/review, and which communicate the results of publicly funded research, is a real impediment to progress.

UPDATE 13th November 2012:

Clearly my gripe about being Locked Out of university benefits – such as access to journals – was timely, because Elsevier has put in place a new scheme called the ‘Postdoc Free Access Passport’. “The program, which is open for applications until December 15, provides unlimited access to books and journals on ScienceDirect for up to six months to young scholars who do not have a research position.” – for more information, read here. Incidentally, this actually looks like it’s an expansion of the free access to reviewers scheme that I commented on in the above post.

[This post was restored from a WayBackWhen archive. It was originally posted to a blog called ‘The Gene Gym” that began life on the Nature Network in 2010, and then moved to Spekrum’s SciLogs platform.]

So why are we interested? Well, if we’re to understand and address the problem of antibiotic resistance, one of the many things we need to do is understand their mechanisms of resistance—how they work. This gene appears very different from any other gene that performs a similar function, and because of this it has been classed into its own ‘family’ of resistance determinant which appears in reviews and textbooks. It has also been screened for, and found, in a notable ‘superbug’, VRSA (the vancomycin resistant big brother of MRSA). The presence of this gene may also have influenced whether or not a patient was given tetracycline because if a genetic screen comes back positive for a tetracycline resistance gene, then you’re not likely to recommend tetracycline (which may have been the preferred drug of choice).

The only problem is, the gene in question is not an antibiotic resistance gene, but we won’t know this until we’ve have spent the summer working on it. Indeed, it won’t be known until the project is inherited as a pet project by a postdoc. The fact is, the gene had already been recognised for what it was over a decade earlier, though this was ever reported. The person who immediately dismissed the gene’s published function all that time ago was in fact the one time PhD supervisor of the postdoc who picked up the project, and a world expert on the family of genes to which our mystery gene belongs, but I’ll come back to that.

My post last week, ‘On publishing negative results…‘, briefly described the issue of positive publication bias in scientific and medical literature, and was a pre-amble to the story of my own experience publishing negative results. So let me now tell you about how I tried, and succeeded, at getting ostensibly negative results published.

Obviously I was the postdoc I describe above, and the project to study this ‘tetracycline resistance gene’ had burned through the summer of an undergraduate and a graduate student, and then through another six months of my time (on and off). It is not uncommon as a postdoc to undertake several smaller projects in parallel to the major project for which the postdoc is employed. Sometimes you win, sometimes you lose. Despite having accumulated plenty of ‘file drawer’ datasets and potential papers over the years, this was one we tried—out of principle—to follow through to publication.

Background

In 1996 a paper from the laboratory of Dr Lolita Daneo-Moore, Temple University PA, reported the isolation of a novel tetracycline resistance gene on a plasmid found within the gut bacterium Enterococcus faecium:

Ridenhour et al. (1996) A novel tetracycline-resistant determinant, tet(U), is encoded on the plasmid pKq10 in Enterococcus faecium. Plasmid 35: 71-80 [DOI]

For information, plasmids are rings of DNA that exist within bacterial cells, but are distinct from the cell’s own DNA. They encode a library of weird and wonderful (though sometimes cryptic) utilities that bacteria can inherit or share between themselves—these utilities often include antibiotic- and antiseptic-resistance, among others.

Whilst I cannot hope to give a full run down of all the details of both this paper and my own paper, in essence Ridenhour’s work represents a classic piece of reductive scientific investigation. Their aim was to find out what made several E. faecium isolates resistant to the antibiotic tetracycline. They honed in on a single small plasmid, ‘pKq10’, present in all the resistant isolates, and having determined the DNA sequence of the plasmid, they found a candidate gene that they called tet(U).  As is the norm, once you have a DNA sequence for something, you can compare this sequence, or the amino acid sequence that the DNA ultimately translates to, with others in databases such as GenBank—to see if anything like it already exists. Based on this sort of computer-based analysis, the authors suggested that it shared some similarity with other types of tetracycline resistance genes, but was different enough to be assigned its own class, Tet(U). Here we use a capitalised ‘Tet(U)’ to describe the protein family, and the lowercase italics ‘tet(U)‘ to describe the gene that encodes it.

It was on this basis, many years later, that our lab decided to figure out how the protein produced by tet(U) actually worked. Our lab has had some success picking apart resistance mechanisms in this way, using a biochemical approach to purify the protein away from the cells and then test its activity. That takes a sentence to say, but took several months to do!

Tetracycline is an antibiotic that targets the protein-making machinery of bacteria. So to study its activity, we can add our purified TetU protein to a small tube that contains an active protein-making soup extracted from bacteria, and see whether its presence protects this protein production from the effects of tetracycline. It didn’t. Nor in fact could we identify any kind of resistance associated with this gene in any other context.

Repeating the original experiments

When I inherited the project, the first thing I did was go back to the original paper and scrutinise both the report, and the DNA sequences they’d submitted to the database. What struck me was that the gene in question, tet(U), looked very familiar. I spent all of 5 minutes double checking the DNA sequence against GenBank and it became clear why. In terms of familiarity, tet(U) was as obvious to me in function as a double-decker bus is to most people. But perhaps the confusion had been that it wasn’t a whole gene I was expecting—it was just the back half. This is because somewhere along the line, when the original lab determined the DNA sequence, two DNA mutations (or sequencing errors) crept in that split a single obvious gene into two, smaller, more cryptic genes (at least to those less familiar with them).

We have to remember that in 1996 sequencing was rather more error prone than it was today. Also, for the sake of ease (and I can’t blame them), the authors had chosen an error prone way to make enough DNA for the sequencing. It’s also true that the GenBank database has had many more submissions in the past 15 years, making the identification of ‘tet(U)‘ more obvious now than back then. It also didn’t help that their bioinformatic analysis stretched the imagination into the patently bizarre—the similarities they reported between tet(U) and other tetracycline resistance genes seemed only possible with wanton desire, rather than empiricism.

So what is tet(U)? It’s actually the back end of a replication gene. This codes for a protein that the plasmid produces so that the plasmid can essentially make more of itself, i.e. replicate. It’s a pretty important protein, and not one that in this family of proteins you would find in two pieces and still be functional. It’s also from the same family of replication proteins with which my former PhD supervisor has a world class expertise.

Given that other researchers and clinical microbiologists were continuing to screen for the presence of tet(U), again potentially informing treatment choices, we felt it prudent to inform the community that it should not be considered a tetracycline resistance determinant. However, with myself and the lab head being self-doubting and meticulous chaps, we thought we’d run through their original experiments, just in case there was something obvious we were missing.

The start of the real problems

The real problem with repeating this work was the fact that Dr Daneo-Moore—senior author of the original paper—is sadly deceased, and the lab and materials have long since dispersed. No-one I contacted had any of the original bacterial isolates, or indeed the plasmid! All we had was the DNA sequence of their plasmid deposited in the database, and the knowledge that tet(U) has cropped up time and again where people have screened for it. By screening, we mean that people are looking for one small section of the gene, rather than trying to understand the genetic context where they find it, i.e. determining whether it’s a distinct gene or in fact part of a (much larger) replication gene. In fact, the saving grace was a recent mass DNA sequencing project that had inadvertently delivered up the sequences I’d need, but I’ll come back to that.

For the time being, I did what I could. I had the whole plasmid synthesised according to the original published DNA sequence, and then dutifully performed all the cuttings and splicings as Ridenhour et al. described, yet saw none of the tetracycline resistance that they did. So this was the point that we decided we’d write a short article to update the literature and have done with it.

The peer review hurdle

I have been a reviewer for six scientific publications for several years, and have performed my duties rigorously and in as scientific a manner possible, looking for sound logic, methodology and substantiated interpretation. The peer review of our paper was a little less than this: one reviewer saw what we were saying, and barring a few stylistic points was happy with it. The other reviewer cost me another five months work.

You see, the thing is, despite pointing at the gene in question and shouting very loudly, “LOOK, IT’S CLEARLY THE BACK END OF A BUS ☞” (i.e. it’s a replication protein, it’s identical to other replication proteins, this is just a small plasmid with one gene and that gene is a replication protein, end of…), the burden of proof was put upon us to disprove that tet(U) was a tetracycline resistance gene. This is of course the essence of Popperism, but we felt we had essentially done this. However, it had been hard to draw a line and say, “tet(U) does not confer tetracycline resistance because it is not a tetracycline resistance gene”, because at the back of my mind I’m thinking, “hmm, does it not confer tetracycline resistance because I am crap and can’t make it do what its supposed to do?”

This, in essence, is the big reviewer ‘fob off’. What researcher has the time to work through so many petty experiments to try to show that the gene isn’t the one thing, all the while ignoring the fairly strong evidence that it’s actually another thing all together?

Sadly, one of the reviewers wasn’t a biochemist, so proceeded to ‘inform’ me (a biochemist) that I couldn’t use the sorts of techniques I’d been using. This is despite my having used them for my entire career, and have been used successfully in biochemistry research for over 15 years, and by others to do just the sort of experiments we’d described. That reviewer also wanted another control experiment, namely to do the same experiments, but on a totally different tetracycline resistance gene, Tet(M). This is a ‘real’ tetracycline resistance gene that the original authors reported as sharing some similarity, but actually it shares no such similarity. Comically, I was also told—in a clairvoyant like manner—that the biochemical approach I’d taken would also “not work” for Tet(M) either. But of course the techniques worked beautifully with Tet(M), and this protein did everything that Tet(U) was supposed to do—but actually didn’t, i.e. confer resistance to tetracycline. So we had at least vindicated the approach!

I was also told that the only proper experiment I could do was to work with the original material used by the lab, and ignore the sequence they had reported in the their paper. So I was being told something that would mean that the hypothesis is now untestable by science, because I could not go back in time and work with exactly whatever it was the original authors were working with. Obviously this is ridiculous—the sequence in their paper is synonymous with tet(U), and is the basis on which it has been accepted in the literature and used to screen for current tet(U) signals in bacteria. If they have failed to include some other important factor, then they have not in fact described the role of tetU (or the true source of resistance) as it stood.

The resubmission

Having performed many more experiments and permutations to generate yet more negative results, I returned to the original tet(U) DNA sequence and compared it against the database again, just for good measure, as it’d been 6 months since my last analysis. Wouldn’t you know, an identical match appeared. I got excited and then inputted the whole sequence of the plasmid, and got an almost identical match with just a few differences. Those differences would be the ones I had predicted would turn an apparent plasmid replication gene into two smaller cryptic genes. Our tet(U) was a mere constituent of a gene labelled as a replication gene on this new matching sequence; this sequence had been submitted to GenBank as part of a partial genome of Enterococcus faecium (the same species as the original host of the plasmid) from a laboratory at University Medical Center Utrecht in the Netherlands.

I just assumed that a plasmid similar to mine had gotten sick of its independent life and inserted itself into the genome of the bacterium—this often happens. I intended to include it as a mere footnote. However, a few days later I found a scientist with a familiar name following me on twitter, Dr Willem van Schaik (@WvSchaik). It took me a while to twig, but realised that coincidentally it was his lab that was doing this sequencing. I sent @WvSchaik a tweet to say that I’d been poring through his sequence data (as you do), but had a question about one contig (the unsorted section of sequence where I’d seen my plasmid). He told me it was in fact a separate plasmid—not part of the genome as I’d thought.

This is where I got more excited and asked @WvSchaik if he wouldn’t mind sending me the bacterial strain this sequence came from. As promised Willem sent the strain. I grew it up, extracted just the plasmid DNA, and found there were actually three plasmids, one of which looked about the right size. A few quick tests later and I had the plasmid, but not any plasmid—it was pKQ10, the original plasmid! It had first been studied 16 years previously in a lab in Pennsylvania, and been lost to science. Now it had turned up in Utrecht, in an Enterococcus strain isolated from an infection in an old man.

And do you know what was interesting about pKQ10? Absolutely nothing. All it does is replicate itself. It is an end unto itself, it exists merely to exist, and does not confer resistance to tetracycline.

And thus:

Caryl et al.(2012) “tet(U)” is not a tetracycline resistance determinant. Antimicrob Agents Chemother 56: 3378-3379 [DOI]

The enterococcal plasmid pKQ10 has been reported to carry a poorly characterized tetracycline resistance determinant designated tet(U). However, in a series of studies intended to further characterize this determinant, we have been unable to substantiate the claim that tet(U) confers resistance to tetracyclines. In line with these results, bioinformatic analysis provides compelling evidence that “tet(U)” is in fact the misannotated 3′ end of a gene encoding a rolling-circle replication initiator (Rep) protein.

If only you knew how careful we had to be with every.choice.of.word throughout the manuscript so that we weren’t accused of being ‘judgemental’.

Thank you to Willem van Schaik (@WvSchaik) at University Medical Center Utrecht for a great little Twitter-based collaboration. I tweet under the handle @jacaryl

N.B. No disrespect is intended, or should be ascribed, to the authors of the original paper. This is simply the iterative nature of science as it should happen, and is a product of progress in molecular and bioinformatic tools, and some stubborn determination to publish negative results.

[This post was restored from a WayBackWhen archive. It was originally posted to a blog called ‘The Gene Gym” that began life on the Nature Network in 2010, and then moved to Spekrum’s SciLogs platform.]

The problem of publication bias in the scientific and medical literature is that positive results get published, and negative results – or those from studies attempting to replicate previous studies – by and large, don’t. There are several problems with this, and with which I’ve had practical experience:

1. Positive results can be the results of flukes, or sheer luck; a consonance of local circumstances that result in a publishable observation. Of course, there will be the necessary internal controls and repetitions that validate a result (one hopes), and these will form the minimal requirements to not be laughed out of the peer review process. However, repeats made within a single laboratory – by the same researchers – can be insufficient, or test too small a component to justify the universality of the conclusion(s) made. Peer reviewers take it ‘on trust’ that the materials and controls being tested are as described, and without contamination.

There is no suggestion that such results are knowingly contrived or maliciously fabricated, more the case that researchers’ naturally enjoy good results, and so they perhaps don’t entertain the necessary rigour once their controls support their conclusion. I don’t necessarily fault this attitude; it is borne of the knowledge that biology is highly variable, with subtle degrees of variation across populations of cells, proteins and DNA. Despite working with the exact same components, it is possible to test something until a new behaviour appears that casts doubt on your previously solid hypothesis. I defy any biochemist or microbiologist who has not encountered a protein that doesn’t behave slightly differently between preparations, or cells that don’t, on occasion, do something a little different from the last three times. Why take the risk? You can always address this in a later paper!

2. Negative results are still results, they can still tell us something new; almost as important as knowing that X causes Y within a given context, is knowing that X doesn’t cause Y within this context. Whilst we might argue that if X causing Y was never known in the first place, publishing the fact that X doesn’t cause Y is going to be a hard sell. However, if X causing Y is already within the literature, the only circumstances where this will be independently verified is when someone not only demonstrates that X causes Y, but also builds upon it; a mere replication would be unlikely to be published. A study that attempted to, but couldn’t, demonstrate that X causes Y, would similarly be unable to publish, unless it managed to show that something else causes Y.

So within the current bias of publishing, the self-correcting nature of science is only possible on results that can be refined and built upon, or debunked whilst also presenting a new positive finding. This leaves a lot of ‘orphan’ results that remain apparently unchallenged within the literature, despite being incorrect.

3. The upshot of this is that the true labourers within science research, grad students, may base aspects of their projects on data that cannot be replicated. This wastes a considerable amount of time because most grad students, being the dutiful and self-doubting scientists they are, are more likely to assume that they did something wrong, rather than the authors of a peer reviewed paper. More painfully, they face a choice, to essentially adopt the study and demonstrate where it went wrong and show what the true result is, therein having something to publish; or continue with their own project, accepting both the loss of time and the fact that they will not be able to publish a cautionary tale.

ON a personal note, I can’t think of a time when I attended a conference and wasn’t either the recipient – or the deliverer – of knowledge that, “So-and-so technique won’t work for that…”, or, “Yeah, we looked at that, we didn’t see anything either…”, to much groaning. It would have been nice had this information been shared in the literature.

4. Perhaps more worringly is when other studies make conclusions based upon the finding of X causing Y, without ever actually verifying this fact; there is no expectation that we should repeat all the work that has ever gone before, thus the fact is we work ‘on trust’ with regards the literature, up to a point. So a lab may have found that that our old friend Y causes Z, and if Z is an undesirable thing, they may ‘reasonably’ conclude that viable targets to address the problem of Z is to target X or Y. Thus the literature branches out, and new investigations become founded upon unrealistic science.

This is of course a gross over-simplification, merely to illustrate a point; granted, scientific research is a little more nuanced than this, but I have my reasons for illustrating these points as I have (see below). It may also be the case that some scientists would choose not to publish negative results, even if they could, in competitive fields where it might be seen as giving competitors a leg-up at your expense. I would add that there have indeed been efforts to catalogue ‘negative’ results, which include databases such as the Negatome, an ambitious effort to list proteins and protein fragments that are known to not interact with each other – useful to remove false positives when looking for proteins that do interact. There have also been various attempts at establishing Journals of Negative Results, with varying degrees of success, though one seemingly active such journal is the Journal of Negative Results in Biomedicine.

My reason for posting this is simply as a pre-amble to my next post, which will be about my experiences with taking on a project that was based on published data that we found to be incorrect, then wasting a lot of time on it, trying to publish it, and having to jump over A LOT of hurdles to actually do so. I will also take the opportunity to address the darker side of peer review that researchers can face. I’ve been meaning to write this story for some time, not least because a Twitter-based conversation made the final part of the project possible, which is as good a reason as any to write about.

Read my next post on ‘My published negative result‘, my experiences of getting negative results published, problematic peer review and the value of Twitter to researchers.

[UPDATE 5th Oct: Think I’ll compile a list of all negative journals here; if anyone knows of any, let me know @jacaryl]

Whilst I appreciate the ethos of these journals, without further investigation I cannot testify as to their quality.

– Journal of Negative Results in Biomedicine
– All Results Journals – Chem, Nano, Biol and Phys [via @CEbikeme]
– Journal of Pharmaceutical Negative Results (!)
– Journal of Negative Results – Ecology and Evolutionary Biology
– Journal of Articles in Support of the Null Hypothesis (via PsychFileDrawer.org)
– The Journal of Spurious Correlations (via PsychFileDrawer.org)

[UPDATE 6th Oct: On the subject of validating the reproducibility of experiments in biomedicine, an editorial in Nature asks some critical questions about the efforts of the Reproducibility Initiative, started by a for-profit start-up based in California. RI aims “to tackle this problem by engaging independent laboratories to repeat experiments and validate results”. Worth a read]

[UPDATE 10th Oct: More discussion: ‘No result is worthless: the value of negative results in science‘, BioMed Central blog]

[This post was restored from a WayBackWhen archive. It was originally posted to a blog called ‘The Gene Gym” that began life on the Nature Network in 2010, and then moved to Spekrum’s SciLogs platform.]

I’m going to take great license to wander around numerous areas that overlap, nudge, cajole and nestle up against the main theme of my blog, which is of course bug and drugs.

So firstly, a brain dump:

A colleague and I once – rather drunkenly – planned a letter to The Lancet [a popular medical journal] in which we describe a means by which one might ‘bank’ a sample of ones faecal matter [shit] (comprising a cross-section of a healthy gut microflora), prior to departing on an exotic holiday, or undergoing antibiotic treatment. The premise was that any insult or injury arising from catching a bout of traveller’s Delhi-Belly, or depletion of the gut flora from chemotherapy, could be abated by having your original gut flora restored from your earlier banked sample. The service would naturally be called, the ‘Shit Bank’.

This would not, however, be a bank to provide samples to others. Faecal transfer is not without hazard, and screening it for a multitude of nasty bugs and viruses would be difficult. Furthermore, your faecal matter is a product of your own environment and diet, so faecal transplantation has always been most effective when the doner is from a close familial/spousal relationship sharing the same resources. The aim is to restore unto you, your very own faecal matter taken during a period of good health, in the same manner that one might bank samples of blood plasma if being stationed in a remote part of the world.

The pressing issue, however, is defining the means by which such faecal matter could be stored. The gut microbiota consists of hundreds of microorganisms, each adapted to specific regions of the gut, and many of which cannot be cultured in the laboratory. Such organisms are referred to as ‘fastidious’, essentially meaning that they’re tough to please. So providing a stable home for them while they’re not, actually, ‘at home’ will prove an issue; will the samples be frozen in a special medium to prevent cell damage? Or perhaps dried down and pelleted? What would the longevity of each major group of microorganisms be? Would there be an opportunity for pathogenic species to take advantage of the change in environment?

I’m not one to let a few minor technical issues come between the Shit Bank and me, and I’m certainly not about to suggest that faecal transplants are a solution to gut health problems; faecal transplantation is not a panacea treatment for all gut ills, and the last thing I want to see is Botox parties being replaced by shit swap parties. However, if your gut is a real mess there could be worse things than kick-starting it with the mother of all probiotic infusions.

[This post was restored from a WayBackWhen archive. It was originally posted to a blog called ‘The Gene Gym” that began life on the Nature Network in 2010, and then moved to Spekrum’s SciLogs platform.]

The trick to identifying if there are latent sleepers operating is to try and re-activate them and get them to reveal themselves. Cue any number of spy stories about false radio signals or targets to lure sleepers into the open.

Curiously, it is a similar strategy that is being used in novel treatments for infections from two disparate areas of chemotherapy, one in the treatment of HIV and the other the treatment of persistent bacterial infections.

In HIV, one of the issues is that the virus can become dormant, they cease to replicate, and remain for all intents and purposes hidden and inactivated in a small pool of resting CD4+ T-cells. The problem with this is that the anti-retroviral drugs we have work by interfering with replication, thus if viruses aren’t actively replicating, they’re not exposed to the drug.

Likewise, in chronic bacterial infections there exists a phenomenon called ‘persistence’. In a bacterial population a sub-population of ‘persister’ cells continue to exist in a stationary or growth-retarded state, regardless of the growth state of the population as a whole. Whilst there are ongoing arguments in the field about what constitutes ‘persistence’ and the genetic or environmental factors that result in such dormancy, it is none the less the case that most antibiotics have been developed to attack actively growing cells. In the absence of growth, the bacteria are simply not effected by the antibiotic.

In both HIV and chronic bacterial infections, removal of the chemotherapy can result in a significant flare-up of the infection as a result of re-activation of the dormant virus/bacteria.

So how do we address these issues? Well, an approach adopted my many HIV/antibiotic research labs is to screen such latent cells/persisters against libraries of different types and shapes of drug molecules. The aim is to identify drug molecules that re-activate the cells, thus forcing the ‘sleepers’ back out into the open where they can be targeted by chemotherapy.

In HIV research circles there has been some work developing two potential approaches. Interleukin-7, a naturally occurring signalling molecule, has shown some signs it can re-activate dormant cells, and thus HIV. The other approach is looking at several histone deacetylase inhibitors, which can reactivate HIV directly. However, one of several major issues has been that it isn’t much good flushing out the sleepers if there are still parts of the body where infected tissues can avoid exposure to such drugs; we’d need to know the extent of such reservoir tissues and how to target them.

In the case of treating bacterial persisters, a recent study demonstrates exactly the sort of prudent, rational approach that can be taken to addressing this issue. The study from the lab of Prof. Jim Collins at Boston University, focussed on targeting both E. coli and S. aureus persister cells with an aminoglycoside antibiotic called gentamicin. Aminoglycosides are a group of drug molecules that interfere with protein synthesis, an essential living process, but they can only work if they can actually get into the cell. It’s well known that the process of aminogylcoside uptake by the cell is energy-driven, however, being metabolically dormant the persister cells have insufficient energy for drug uptake and so gentamicin has only weak activity.

Prof. Collins team found that treating persister cells with certain sugars (glucose, fructose, and mannitol) induced rapid-killing when gentamicin was present, and so seemingly the idea of metabolically stimulating such dormant cells hold true. The addition of sugars doesn’t exactly turn these cells into native active cells, because this treatment only works to enable aminoglycoside-mediated killing. Other classes of antibiotics still had no effect. Sugars such as glucose, fructose and mannitol enter a cell’s metabolism at the head of a process that generates high-energy molecules. Whilst this energisation is enough to enable aminoglycoside uptake, it’s not enough to get all the cellular processes ticking over, hence the lack of reaction to antibiotic drug classes that target other cellular processes.

What is interesting is that the process works for both Gram-negative bacteria (such as E. coli) and Gram-positive bacteria ( S. aureus ), both representing a broad divide in the bacterial world, though in S. aureus only the fructose, rather than glucose and mannitol, resulted in rapid killing by gentamicin.

In our spy novel we might say that the enemy sleeper agent is being lured out into the open by a sugar coated package, the contents of which spell the agent’s death. But certainly approaches such as these can only benefit the increasingly complex requirements of antibacterial chemotherapy that are required to treat persistent infections.

Ref.

Allison KR, Brynildsen MP and Collins JJ. (2011) Metabolite-enabled eradication of bacterial persisters by aminoglycosides. Nature 473: 216-220. doi: 10.1038/nature10069
[ pdf ]

Silva RF, Mendonça SCM, Carvalho LM, Reis AM, Gordo I, et al. 2011 Pervasive Sign Epistasis between Conjugative Plasmids and Drug-Resistance Chromosomal Mutations. PLoS Genet 7(7): e1002181. doi: “10.1371/journal.pgen.1002181 “:http://dx.doi.org/10.1371/journal.pgen.1002181

Following this, I have been quoted in a NewScientist Health News item, and as I don’t feel my response is quite in the context I gave it, I thought I would give a more detailed account. I spoke to NewScientist last Wednesday (the paper in question wasn’t due to be published until the following day), but I was told that in the study the authors had observed that antibiotic resistance can have a positive effect on bacterial fitness, even in the absence of selection. I was asked whether this was a surprise to me, and more generally about research in bacterial fitness. What I perhaps should have done was ask specifically whether the paper was still embargoed and whether I could have more particulars of the study, because I could not have anticipated the particular nature of the study in question.

The paper describes an observation that the fitness costs of chromosomal mutations conferring resistance to antibiotics, whilst initially deleterious, can become beneficial through the acquisition of a transferable antibiotic resistant plasmid. The outcome of this might be cause for concern given the widespread occurrence of conjugative resistance plasmids in clinical strains; the idea that these may be stabilising otherwise costly resistance mutations is worrying.

We know from many studies that some mutations that give rise to antibiotic resistance come at a cost to bacteria. A bacterium may for example overcome an antibiotic by having a mutation in the key protein that the antibiotic targets, but the mutation may also make that protein less capable at its day-to-day function, and this presents a fitness cost to the cell. Obviously while there is antibiotic around, such mutations bring a strong fitness advantage, but when the antibiotic is no longer around, and these cells are forced to compete with their antibiotic-sensitive brethren, they can become a fitness cost.

The phenomenon in which the effect of one set of genes (say those encoded on a conjugative plasmid) may have a synergistic effect on another set of genes (i.e. those on the chromosome) that results in a more beneficial outcome is called ‘positive epistasis’. Whilst I’m familiar with this phenomenon more generally in bacterial metabolism, as it has been documented by the inimitable Richard Lenski over the years, I would be hard pressed to cite examples where two negative fitness costs (one chromosomal, and one plasmid-borne) might cancel each other out, or indeed even result in a beneficial fitness advantage that is more than the sum of its constituent costs.

However, as I often quote, Crichton’s Jurassic Park character Dr Ian Malcolm would say, ‘Life finds a way…” Indeed, bacteria that encounter a fitness cost may under go ’mitigating’ mutations that alleviate the fitness costs of the original mutation, restoring their fitness to that of their non-resistant brethren. This way the bacteria get to remain resistant at no extra cost, kind of like having your cake, and eating it too. It was this much that I described in my conversation with NewScientist. I have previously described the means by which bacteria may mitigate fitness costs (second half of post).

The study in question is more observational than mechanistic, so we are not clear why it is that acquisition of a conjugative resistance plasmid can be beneficial to cells already encoding chromosomal resistance to an antibiotic. This will likely have to await further investigation, however, the question inevitably turns to how to address this issue. As I mentioned to NewScientist, one aspect of my research is to identify those antibiotics against which resistance imposes the greatest fitness costs, the idea being that when resistance occurs, it may persist for less time once the use of that antibiotic is curtailed. However, this is not a one-stop solution, it would form one of many small changes in practise to prevent antibiotic persistence.

I don’t disagree with Dionisio’s idea of targeting the plasmids, and looking for ways to inhibit plasmid conjugation and replication, thus addressing one of the major causes of antibiotic resistance spread. Indeed, such approaches have been reviewed previously (Williams & Hergenrother, 2008, Exposing plasmids as the Achilles’ heel of drug-resistant bacteria. Current Opinion in Chemical Biology 12: 389-399 doi: 10.1016/j.cbpa.2008.06.015), and some interesting chemical agents found.

In fact, I wonder if the presence of conjugative plasmids reduce the fitness costs of chromosomal mutations to the extent that the typical mitigating mutations seen to stabilise other chromsomal mutations don’t occur. This at least presents the opportunity that a combined attack on the plasmid and the cell may be successful in eradicating particular resistance determinants before they become independently stable.

I’m glad that NewScientist took an interest in this paper as I think we need to keep the issue of antibiotic resistance, and what we learn about the associated problems and the solutions, in the public eye. I’m also open for comments on this area, but perhaps next time I will clarify what the interviewer has understood from my responses.