Open source software tool for the week: AXIOME: automated exploration of microbial diversity

Just found this paper via Google Scholar Updates:
GigaScience | Abstract | AXIOME: automated exploration of microbial diversity

From Josh Neufeld's lab this paper describes a series of tools for automation of microbial diversity analyses.

Abstract:
BackgroundAlthough high-throughput sequencing of small subunit rRNA genes has revolutionized our understanding of microbial ecosystems, these technologies generate data at depths that benefit from automated analysis. Here we present AXIOME (Automation, eXtension, and Integration Of Microbial Ecology), a highly flexible and extensible management tool for popular microbial ecology analysis packages that promotes reproducibility and customization in microbial research.
FindingsAXIOME streamlines and manages analysis of small subunit (SSU) rRNA marker data in QIIME and mothur. AXIOME also implements features including the PAired-eND Assembler for Illumina sequences (PANDAseq), non-negative matrix factorization (NMF), multi-response permutation procedures (MRPP), exploring and recovering phylogenetic novelty (SSUnique) and indicator species analysis. AXIOME has a companion graphical user interface (GUI) and is designed to be easily extended to facilitate customized research workflows.
ConclusionsAXIOME is an actively developed, open source project written in Vala and available from GitHub (http://neufeld.github.com/axiome) and as a Debian package. Axiometic, a GUI companion tool is also freely available (http://neufeld.github.com/axiometic). Given that data analysis has become an important bottleneck for microbial ecology studies, the development of user-friendly computational tools remains a high priority. AXIOME represents an important step in this direction by automating multi-step bioinformatic analyses and enabling the customization of procedures to suit the diverse research needs of the microbial ecology community.

Workflows like this are what many many people need.  I note - I have not used this yet but it looks promising.  It has some parallels to the WATERS workflow package that came from my lab a few years ago (see more about it here: http://phylogenomics.wordpress.com/software/waters/.)  Alas WATERS is no longer being actively developed.  Anyway - AXIOME has additional features and certainly seems like it would be useful to many people.

Guest post from Ashley Bateman on "Full contact microbes" - Roller Derby

A special guest post from Ashley Bateman.

 

Roller derby players share their skin microbes during play

Single-celled organisms are intimately associated with multicellular organisms across the tree of life, and human beings are no exception. Making up 90% of our cellular composition, these invisible passengers (our microbiome) contribute to our health and well-being in crucial ways, including aiding our digestion, the education of our immune system, and resistance to pathogens. Despite this importance, we still lack a fundamental understanding of where our host-associated microbes actually come from. We know that infants are born practically sterile; early-life events such as birth mode can contribute to the types of microbial species found on an individual, but these events cannot adequately explain the majority of spatiotemporal variation observed over a host’s lifetime. To be able to accurately describe the processes that drive host-associated microbial community dynamics, we must have an informed understanding of the role of dispersal in structuring host-associated microbial communities.

Where do they come from? How do they get there? Do these changes (if any) last?

The Green Lab at the University of Oregon-Eugene attempted to answer some of these questions in our latest publication “Significant changes in the skin microbiome mediated by the sport of Roller Derby”, released today by PeerJ. We decided to use Roller Derby as a model system to investigate the role of contact in dispersing skin microbial communities between hosts. We have known for a long time that pathogens can be transmitted via direct contact; could not our commensal microbial communities be shared in this way?

We swabbed the upper arms (a frequent contact point between players during a bout) of players belonging to 3 geographically distinct derby teams and characterized the skin microbiome of each player using 16s rRNA gene Illumina sequencing. We found that each team’s upper arm microbiome was significantly different from one another before play, and that this difference decreased after bouts were played. Not only did teams’ skin microbiomes become less different from one another after play, but the differences were driven in part “by the presence of unique indicator taxa that are commonly associated with human skin, gut, mouth, and respiratory tract.” There were also environmental bacteria associated with soil and plants found in the skin samples.

Although we weren’t able to show a direct link between contact and transfer of specific microbial taxa, the best explanation of the data seems to be that contact between these players during a one-hour bout effectively resulted in homogenization of their upper arm skin microbiomes.

So much yet to explore! As a 2^nd year graduate student in the Green Lab I hope to address some of the questions that the Roller Derby paper has brought to our attention. My dissertation research is gearing up to understand the role of dispersal on our skin microbiome. Are some skin sites more amenable to changes than others? Can we pick up host-associated microbes not just from other individuals, but from objects that other individuals have touched? Can we pick up non-host-associated microbes? If we can pick them up, how long do they stick around? How do they participate in the functions attributed to the skin microbiome?

Hope to keep up the fantastic momentum that has been launched by this latest publication – if you have any thoughts or comments, feel free to contact me at abateman@uoregon.edu, or via Twitter: @microbesrock

And you can check out a stop-motion video I made on the skin microbiome here:

Interesting paper on strategy to use PCR to simultaneously characterize eukaryotic, bacterial and archaeal microbes

Interesting new paper in PLOS One: PLOS ONE: Simultaneous Amplicon Sequencing to Explore Co-Occurrence Patterns of Bacterial, Archaeal and Eukaryotic Microorganisms in Rumen Microbial Communities

Full citation:  Kittelmann S, Seedorf H, Walters WA, Clemente JC, Knight R, et al. (2013) Simultaneous Amplicon Sequencing to Explore Co-Occurrence Patterns of Bacterial, Archaeal and Eukaryotic Microorganisms in Rumen Microbial Communities. PLoS ONE 8(2): e47879. doi:10.1371/journal.pone.0047879

Basically, the paper describes the development and use of a PCR strategy to simultaneously characterize eukaryotic, bacterial and archaeal microbes from samples.

Primers used are summarized in Table 2

The strategy they employ attempt to correct for differences in amplification differences between the different amplicons which should therefore allow better normalization of relative abundance estimates.  See results in Figure 2.

Definitely worth a look.

Correcting for rRNA copy # in qPCR experiments

Asked this question on Twitter and thought I would share answers here via Storify.  I am putting it below the fold to allow people to avoid the Storify embed if they want to.

Interesting new #PLOS One paper on study design in rRNA surveys

Interesting new paper in PLoS One:  PLOS ONE: Taxonomic Classification of Bacterial 16S rRNA Genes Using Short Sequencing Reads: Evaluation of Effective Study Designs

Abstract: Massively parallel high throughput sequencing technologies allow us to interrogate the microbial composition of biological samples at unprecedented resolution. The typical approach is to perform high-throughout sequencing of 16S rRNA genes, which are then taxonomically classified based on similarity to known sequences in existing databases. Current technologies cause a predicament though, because although they enable deep coverage of samples, they are limited in the length of sequence they can produce. As a result, high-throughout studies of microbial communities often do not sequence the entire 16S rRNA gene. The challenge is to obtain reliable representation of bacterial communities through taxonomic classification of short 16S rRNA gene sequences. In this study we explored properties of different study designs and developed specific recommendations for effective use of short-read sequencing technologies for the purpose of interrogating bacterial communities, with a focus on classification using naïve Bayesian classifiers. To assess precision and coverage of each design, we used a collection of ~8,500 manually curated 16S rRNA gene sequences from cultured bacteria and a set of over one million bacterial 16S rRNA gene sequences retrieved from environmental samples, respectively. We also tested different configurations of taxonomic classification approaches using short read sequencing data, and provide recommendations for optimal choice of the relevant parameters. We conclude that with a judicious selection of the sequenced region and the corresponding choice of a suitable training set for taxonomic classification, it is possible to explore bacterial communities at great depth using current technologies, with only a minimal loss of taxonomic resolution.

Not sure I like everything in the paper.  For example, they focus on naive Bayesian classification methods ... when (of course) I prefer phylogenetic methods.  But that is a small issue.  Overall there is a lot of useful detail in here about rRNA based taxonomic studies.  I note - some of this probably applies to metagenomic studies as well ... perhaps this group will do a comparable analysis of metagenomics next?

Mizrahi-Man O, Davenport ER, Gilad Y (2013) Taxonomic Classification of Bacterial 16S rRNA Genes Using Short Sequencing Reads: Evaluation of Effective Study Designs. PLoS ONE 8(1): e53608. doi:10.1371/journal.pone.0053608

RIP Carl Woese: Collecting posts / notes / other information about my main science hero here

My tribute to Carl Woese 12/30/12

Sadly, Carl Woese has passed away.  I am collecting some links and posts about him here in his memory.  He was without a doubt the person who most influenced my career as a scientist.

News stories about Woese's passing

• Champaign/Urbana News-Gazette Visionary UI biologist Carl Woese, 84, dies
• U. of I. News: Renowned microbiologist Carl R. Woese dies at age 84
• University professor of microbiology dies at 84 years old
• Chicago Sun Times: Revolutionary U of I biologist Carl R. Woese dies
• NY Times: Carl Woese Dies at 84; Discovered Life's 'Third Domain'
• Wired: Carl Woese, (R)evolutionary Biologist
• Huff Post: Evolution Scientist Carl Woese Dies: 'The Most Important Evolution Scientist of the 20th Century'
• Carl Woese Dies | The Daily Scan | GenomeWeb
• Chicago Tribune: Carl R. Woese, microbiologist and professor at U. of I., 1928-2012

Some of my posts about Woese

• January 2013 Reading in detail Carl Woese's 1998 "Manifesto on Microbial Genomics" for the first time ...
• April 2010 Most important paper ever in microbiology? Woese & Fox, 1977, discovery of archaea
• October 2010: Here's hoping molecular classification/systematics of cultured & uncultured microbes wins #NobelPrize in medicine ...
• September 2012: Some arguments for why Carl Woese (and probably Norm Pace) deserves a Nobel Prize
• Notes from a talk by Woese in 1993
• Evolution pumpkins - from Darwin to Woese

Woese Tree of Life pumpkin (by J. Eisen)

Storification of Tweets and other posts about his passing

Other posts worth reading about Woese's passing

• So long, and thanks for all the Archaea » Pharyngula
• Prof. Carl Woese – 1928-2012 | C.W. Schadt | Microbial Ecology Lab
• A Sad Post: RIP Carl Woese! - Creatures Great AND Small ...
• T. taxus: Carl Woese 1928-2012
• Carl Woese - NASA Astrobiology: Life in the Universe
• Honoring a scientific revolutionary from Rob Dunn
• In Memoriam Carl Woese from Karl Fogel's blog
• Carl Woese dead at 84 - The Panda's Thumb
• RIP Carl Woese: Evolutionary Pioneer from the DailyKos
• From Larry Smarr
• Reposting of a Woese interview in LaborJournal.De
• El árbol que plantó Carl R. Woese
• Archaea Are More Wonderful Than You Know
• A sad loss for Microbiology
• Curiosidades de la Microbiología: Carl Woese 1928-2012
• The new biology of Carl Woese

Some videos with Woese 

Miscellaneous

My graduate student Russell Neches used a laser to etch a picture of Carl Woese on a piece of toast.

More about Carl Woese

• Carl Woese - Wikipedia, the free encyclopedia
• Carl R Woese - The School of Molecular and Cellular Biology ...
• Phenomenal history / discussion of Woese's work: Phylogeny and beyond: Scientific, historical, and conceptual significance of the first tree of life
• 2007 Science article by Virginia Morell

Papers by Carl Woese is a group in Biological Sciences on Mendeley.

People not Projects: the Moore Foundation continues to revolutionize marine microbiology w/ its Investigator program

People not Projects.

It is such a simple concept.  But it is so powerful.  I first became aware of this idea as it relates to funding scientific research in regard to the Howard Hughes Medical Institute's Investigator program.  Their approach (along with a decent chunk of money) has helped revolutionize biomedical science.  And thus I was personally thrilled to see the introduction of this concept in the area of Marine Microbiology a few years back with the Gordon and Betty Moore Foundation's "Marine Microbiology Initiative Investigator" program.  Launched in 2004 it helped revolutionize marine microbiology studies in the same way HHMI's investigator program revolutionized biomedical studies.

The first GBMF MMI Investigator program ran from 2004 -2012. And the people supported were pretty darn special:

• Dr. E. Virginia Armbrust, University of Washington  
• Dr. Sallie W. Chisholm, Massachusetts Institute of Technology  
• Dr. Edward F. DeLong, Massachusetts Institute of Technology  
• Dr. Stephen J. Giovannoni, Oregon State University  
• Dr. David M. Karl, University of Hawaii  
• Dr. Nicole King, UC Berkeley  
• Dr. Jennifer Hughes Martiny, UC Irvine  
• Dr. Mary Ann Moran, University of Georgia  
• Dr. Victoria Orphan, California Institute of Technology  
• Dr. Forest Rohwer, San Diego State University  
• Dr. Alexandra Z. Worden, Monterey Bay Aquarium Research Institute  
• Dr. Jonathan Zehr, UC Santa Cruz  

Now I am I suppose a little biased in this because at the same time GBMF launched this program they also put a bunch of money into the general area of Marine Microbiology and I have been the recipient of some of that money.  For example, I got a small amount of money as part of the GBMF Funded work at the J. Craig Venter Institute on the Sargasso Sea and Global Ocean Sampling metagenomic sequencing projects and also had a subcontract from UCSD/JCVI to do some work as part of the "CAMERA" metagenomic database project.  I ended up being a coauthor on a diverse collection of papers associated with these projects including Sargasso metagenome and this review, and GOS1, GOS2 and my stalking the 4th domain paper.

I am also a bit biased in that I have worked with many of the people on the initial MMI Investigator list some before, some after the awards including papers with Jen Martiny, Ed Delong, Alex Worden and Ginger Armbrust, and Mary Ann Moran.

But perhaps most relevant in terms of possible bias towards the Gordon and Betty Moore Foundation is that in 2007 my lab received funds through the MMI program for a collaborative project with Jessica Green and Katie Pollard for our "iSEEM" project on "Integrating Statistical, Ecological and Evolutionary analyses of Metagenomic Data" (see http://iseem.org) which was one of the most successful collaborations in which I have ever been involved.  This project produced something like a dozen papers and many major new developments in analyses of metagenomic data including 16S copy correction, sifting families, microbeDB, PD of metagenomes, WATERs, BioTorrents, AMPHORA. and STAP.  This project just ended but Katie Pollard and I just got additional funds from GBMF to continue related work.

So sure - I am biased.  But the program is simply great.  In the eight years since the initial grants the Gordon and Betty Moore Foundation has helped revolutionize marine microbiology.  And a lot of this came from the Investigator program and it's emphasis on people not projects.  I note - the Moore Foundation has clearly decided that this "people not projects" concept is a good one.  A few years ago they partnered with HHMI to launch a Plant Sciences Investigator Program  which I wrote about here.

It was thus with great excitement that I saw the call for applications for the second round of the MMI Investigator program.  I certainly pondered applying.  But for many reasons I decided not to.  And today the winners of this competition have been announced and, well, it is an very impressive crew:

• Andrew Allen, J. Craig Venter Institute
• Ginger Armbrust, University of Washington
• Kay Bidle, Rutgers University
• Edward DeLong, Massachusetts Institute of Technology
• Curtis Deutsch, University of California-Los Angeles
• Nicole Dubilier, Max Planck Institute for Marine Microbiology
• Michael Follows, Massachusetts Institute of Technology
• Jed Fuhrman, University of Southern California
• David Karl, University of Hawaii
• Victoria Orphan, California Institute of Technology
• Ann Pearson, Harvard University
• Forest Rohwer, San Diego State University
• Mak Saito, Woods Hole Oceanographic Institution
• Roman Stocker, Massachusetts Institute of Technology
• Matthew Sullivan, University of Arizona
• Alexandra Worden, Monterey Bay Aquarium Research Institute

Some of the same crowd as the previous round.  Some new people.  Some people not there from the previous round.  All of them are rock stars in their areas especially if one takes into account how senior they are (the more junior people are stars in development).  And all have done groundbreaking work in various areas relating to marine microbiology.  The organisms covered here run the gamut including viruses, bacteria, archaea, and microbial eukaryotes.  The areas of focus covered range from biogeochemistry to ecosystem modeling with everything in between.  It really is an impressive group. Delong pioneered metagenomics and helped launch studies of uncultured microbes in the oceans.  Karl has led the Hawaii Ocean Time series and done other brilliant work.  Sullivan and Rohwer and pushing the frontiers of viral studies in the oceans.  Allen, Armbrust, and Worden are among the leaders in genomic studies of microbial eukaryotes in the marine environment.   Dubilier, Bidle, Fuhrman and Follows Stocker (double listed Follows in original post ...) - though they focus on very different aspects of marine microbes - are helping lead the charge in understanding interactions across the domains of life in the marine environment.  Orphan, Saito, Deutsch, Follows and Pearson are on the cutting edge of biogeochemical studies and trying to link experimental studies of microbes to biogeochemistry of oceans.

The great thing about the "people not projects" concept is that the people funded here get to follow their own path.  They are not going to be constrained by the complications and sometime idiocy of the grant review process.  They in essence get to do whatever they want.  Freedom to follow their noses.  Or their guts.  Or whatever.  It is a refreshing concept and as mentioned above has been revolutionary in various areas of science.  There has been a slow but steady spread of the "people not projects" concept to various federal agencies too but it seems to be more of a private foundation type of strategy.  Federal Agencies are so risk averse in funding that this type of concept does not work well there.  I wish there was more.  But I am at least thankful for what HHMI and GBMF and Wellcome and Sloan and other private groups are doing in this regard.  Now - sure - all of these private foundations do not do everything perfectly.  They have blunders here and there like everyone else.  But without a doubt I think we need more of the People not Projects concept.

Oh - and another good thing.  GBMF is quite a big supporter of Open Science in it's various guises.  So one can expect much of the data, software, and papers from their funding to be widely and openly available.   

It is a grand time to be doing microbiology largely due to revolutions in technology and also to changes in the way we view microbes on the planet.  It is an even grander time to be doing marine microbiology due to the dedication of the Gordon and Betty Moore Foundation to this important topic.  

Referring to 16S surveys as "metagenomics" is misleading and annoying #badomics #OmicMimicry

Aargh.  I am a big fan if of ribosomal RNA based surveys of microbial diversity.  Been doing them for 20+ years and still continue to - even though I have moved on to more genomic/metagenomic based studies.  But it drives me crazy to see rRNA surveys now being called "metagenomics".

Here are some examples of cases where rRNA surveys are referred to as metagenomics:

• Deep 16S rRNA metagenomics and quantitative PCR analyses of the premature infant fecal microbiota ... - Wow -- rRNA as metagenomics even made it into the title here
• Paper: 16S rRNA metagenomics-based survey of oral biofilms in obese children... Poster Abstract
• Gastroenterology & Endoscopy News - Studies Link Composition of ... News Story
• Metagenomic study of the oral microbiota by Illumina high-throughput sequencing - paper is only about rRNA sequencing, not metagenomics
• Dr. Dag Harmsen publishes first 16S metagenomic study on the Ion PGM Sequencing ... a Youtube video highlighting a PLoS One paper
• Bacterial Community Shift in Treated Periodontitis Patients Revealed by Ion Torrent 16S rRNA Gene Amplicon Sequencing - PLOS One paper from the video above
• EUREKA GENOMICS | 16S metagenomic analysis service - a company pushing their services
• A Metagenomic Approach to Characterization of the Vaginal Microbiome Signature in Pregnancy ... PLoS One paper

I found these examples in about five minutes of googling.  I am sure there are many many more.  

Why does this drive me crazy?  Because rRNA surveys focus on a single gene.  They are not gnomicy in any way.  Thus it is misleading to refer to rRNA surveys as "metagenomics".  Why do people do this?  I think it is pretty simple.  Genomics and metagenomics are "hot" topics.  To call what one is doing "metagenomics" makes it sound special.  Well, just like adding an "omic" suffix does not make ones work genomics - falsely labeling work as some kind of "omics" also does not make it genomics.

Enough of this.  If you are doing rRNA surveys of microbial communities - great - I love them.  But do not refer to this work as metagenomics.  If you do, you are being misleading, either accidentally or on purpose.    So I think I need a new category of #badomics - "Omic Mimicry" or something like that ...

------------------------------

Note - this post was spurred on by a Twitter conversation - which is captured below (note - I am certain I have complained about this before but cannot find a record of it ...)

Nice review on HiSeq/MiSeq rRNA sequencing from Caporaso et al #microbes

Quick post -- nice review worth checking out: The ISME Journal - Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms

from Caporaso JG, Lauber CL, Walters WA, Berg-Lyons D, Huntley J, Fierer N, Owens SM, Betley J, Fraser L, Bauer M, Gormley N, Gilbert JA, Smith G, Knight R.

A key part of the paper, with highlighting from me:-

These observations, in agreement with studies that have addressed this question directly (Kuczynski et al., 2010), suggest that increasing the sequencing depth is not likely to provide additional insight into questions of beta diversity, and we therefore argue that (for questions of beta diversity in particular) the decreased cost of sequencing should be applied to study microbial systems using many more samples, for example, in dense temporal or spatial analyses, rather than with many more sequences per sample.  Of course, if the objective is to identify taxa that are very rare in communities, deeper sequencing will be advantageous. Additionally we note that while as few as 10 sequences per sample may be useful for differentiating very different environment types (for example, soil and feces), as environments become more similar (for example, two soil samples of different pH) more sequences will be required to differentiate them.

Journal club today on bacteria in toilets - posting some notes here

I am heading a journal club discussion today of the following paper: PLoS ONE: Microbial Biogeography of Public Restroom Surfaces

I am going to use this page/post to put up some notes for the discussion.  Fortunately I have a good guide in this - Rob Dunn wrote a nice commentary/review for Scientific American blogs: Public bathrooms house thousands of kinds of bacteria

Stay tuned/come back to this page as I will be posting some more notes. Any suggestions for other things to look at/discuss would be welcome.

Notes (I note - I am copying much of the text from the paper not rewriting it.)

Coming Monday at #UCDavis "The Infant Gut Microbiome: Prebiotics, Probiotics, & Establishment"

Just a little announcement here.  There is a symposium tomorrow at UC Davis organized by a undergraduates in the CLIMB program.  CLIMB stands for "Collaborative Learning at the Interface of Mathematics and Biology (CLIMB)" and is a program that emphasizes hands-on training using mathematics and computation to answer state-of-the-art questions in biology.  A select group of undergraduates participate in the program and this summer the students had to do some sort of modelling project.  Somehow I managed to convince them to do work on human gut microbes.  And they have done a remarkable job.

As part of their summer work, they organized a symposium on the topic and their symposium takes place tomorrow.  Details are below.

The Infant Gut Microbiome: Prebiotics, Probiotics, & Establishment

Monday, 12 September 2011, 9am-4pm

Life Sciences 1022

UC Davis

9:00-9:10 Introduction

9:10-9:40 Jonathan Eisen, UC Davis

“DNA and the hidden world of microbes”

9:40-10:40 Mark Underwood, UC Davis

“Dysbiosis and necrotizing enterocolitis”

10:40-10:50 break

10:50-11:50 Ruth Ley, Cornell University

“Host-microbial interactions and metabolic syndrome”

11:50-12:00 general discussion

12:00-1:00 lunch

1:00-2:00 CLIMB 2010 cohort

“Breast milk metabolism and bacterial coexistence in the infant microbiome”

2:00-2:10 break

2:10-3:10 David Relman, Stanford University

“Early days: assembly of the human gut microbiome during childhood"

3:10-3:40 Bruce German, UC Davis

3:40-4:00 next steps

The only major issue for me is I am losing my voice.  So we will see how this goes.  Though I note I have gotten some very sage advice on how to treat my voice problem via the magic of twitter.  If I do not collapse I will also be tweeting/posting about the other talks during the day.

The story behind the story of my new #PLoSOne paper on "Stalking the fourth domain of life" #metagenomics #fb

Well, here goes.

This is a post about a paper that has been a long long time coming. Today, a paper of mine is being published in PLoS One. The paper is titled "Stalking the Fourth Domain in Metagenomic Data: Searching for, Discovering, and Interpreting Novel, Deep Branches in Marker Gene Phylogenetic Trees" and is available at http://dx.plos.org/10.1371/journal.pone.0018011. (or if that link does not work you can get a copy here). This paper represents something I started a long time ago and I am going to try to describe the story behind the paper here.

I note - we are not doing a press release for the paper, for a few reasons. But one of them is that, well, I am starting to hate press releases. So I guess this is kind of my press release. But this will be a bit longer than most press releases. I note - my key fear here is that somehow in my communications with the press or in our text in the paper or in this post I will overstate our findings. Here is the punchline - we found some very phylogenetically novel forms of phylogenetic marker genes in metagenomic data. We do not have a conclusive explanation for the origin of these sequences. They may be from novel viruses. The They may be ancient paralogs of the marker genes. Or they may be from a new branch of cellular organisms in the tree of life, distinct from bacteria, archaea or eukaryotes. I think most likely they are from novel viruses. But we just don't know.

UPDATE: Am posting some links here to news stories/blogs about our paper

• Glimpses of the Fourth Domain? by Carl Zimmer at his always stellar "The Loom" 
• Biology's 'dark matter' hints at fourth domain of life by Colin Barras at the New Scientist 
• 21st century science publishing will be multilevel and multimedia by PZ Myers 
• German Radio Story
• Chinese news report
• PLoS One Blog Pick of the Month for March 2011 by Brian Mossop 
• Brazilian news story from @info
• There might be at least one other, previously hidden, domain of life by Francis Martin 
• Pronađen potpuno novi "domen" života
• Craig Venter's study of marine DNA finds new branches on the tree ... Telegraph article #1 
• Scientist finds a whole new 'domain' of life - Telegraph article #2 
• Scientist stumbles on new class of living things - Zee News 
• A new domain of life: Plenty more bugs in the sea from The Economist 
• DNA Analysis Hints At a Fourth Domain of Life - Slashdot
• Check out the video story from Newsy.Com

Norman R. Pace visit to #UCDavis; discussing microbiology of the built environment #microBEnet

Norman R. Pace, from UC Boulder, gave a talk at UC Davis last week about microbial diversity.  In his talk he discussed some of his recent Sloan Foundation funded work on “microbiology of the built environment” including studies of shower heads, indoor swimming pools, water supplies, and hospitals.

Pace is one of the pioneers of DNA based studies of microbes in the environment.  His initial work on studies of ribosomal RNA from uncultured organisms (started more than 20 years ago) helped launch the field.  For more information on his work see his lab page here

If you are interested in the microbes that are found in showerheads, his PNAS paper on this (which can be found here) on this from 2 years ago got a lot of press.  See for example this Science Friday
and this New York Times article by Nicholas Wade.

Pace was at UC Davis as part of the Storer Major Issues in Modern Biology Lecture Series.

I note, I have written about Pace before a few times including this:
Here's hoping molecular classification/systematics of cultured & uncultured microbes wins #NobelPrize in medicine

I note we have a new project as part of this Sloan program to facilitate communication and networking and sharing information as part of this project.  My lab is creating something called "microBEnet" - the microbiology of the built environment network.  We are just getting our real site up and running.  For now you can find out some information at a temporary page http://microbenet.blogspot.com/

Phylogeny rules:

I am a coauthor on a new paper in PLoS Computational Biology I thought I would promote here.  The full citation for the paper is:

PhylOTU: A High-Throughput Procedure Quantifies Microbial Community Diversity and Resolves Novel Taxa from Metagenomic Data (doi:10.1371/journal.pcbi.1001061). 

The paper discusses a new software program "phylOTU" which is for phylogenetic-based identification of "operational taxonomic units", which are also known as OTUs.   What are OTUs?  They are basically clusters of closely related sequences that are used to represent something akin to a species.  OTUs are used a lot in environmental microbiology b/c one key way to study microbes in the environment is through extraction and sequencing of DNA.  Traditionally this has been done through PCR amplification and sequencing of one particular gene (ss-rRNA).  Now it is also being done through random sequencing of all DNA from environmental samples (so called metagenomics).

Anyway - the paper is (of course) fully open access and you can read it for more detail.  Just thought I would post a little here about it.  The paper / project was led by Tom Sharpton, a post doc in Katie Pollard's lab at UCSF working on a collaborative project between Katie's lab, my lab, and Jessica Green's lab at U. Oregon (and recently Martin Wu's new lab at U. Virginia - he was in my lab previously).  This collaborative project even has a name "iSEEM" which stands for integrating statistical, evolutionary and ecological approaches to metagenomics.  This project has been generously supported by the Gordon and Betty Moore Foundation (via a grant for which I am PI).

Some little tidbits of possible interest about the project

• I really wanted to program to be called POTUS, but I guess I lost out ...
• You can get the code here: https://github.com/sharpton/PhylOTU 
• You can also get code/data here: http://www.biotorrents.net

Sharpton, T., Riesenfeld, S., Kembel, S., Ladau, J., O'Dwyer, J., Green, J., Eisen, J., & Pollard, K. (2011). PhylOTU: A High-Throughput Procedure Quantifies Microbial Community Diversity and Resolves Novel Taxa from Metagenomic Data PLoS Computational Biology, 7 (1) DOI: 10.1371/journal.pcbi.1001061

Here's hoping molecular classification/systematics of cultured & uncultured microbes wins #NobelPrize in medicine

From Wu et al. 2009. A phylogeny driven genomic encyclopedia of bacteria and archaea. Nature 462, 1056-1060 doi:10.1038/nature08656  http://bit.ly/8Y8xea

Well, I am always hopeful.  Every year when the Nobel Prizes come around I am alway hoping that one of them goes to someone involved in studying microbial diversity in some way.  And really, there is a potential Nobel Prize in Physiology or Medicine out there in this area.  Sure they do not give out a Nobel in biology, or evolution or ecology.  But I think a good argument could be made for giving out a Nobel Prize in Physiology or Medicine to those who have worked on molecular systematics of cultured and uncultured microbes.

Why should this attract the attention of those giving out the Nobel Prizes?  Well, without molecular systematics of microbes we would be completely lost in a sea of microbial diversity.  And with such molecular systematics we can not only make much more sense out of the biology of cultured organisms, but we can go to environments and determine who is out there by sampling their genes.  And this type of work has undoubtedly revolutionized medicine, from determining what antibiotics are most likely to be useful in infections, to tracking emerging infectious diseases, to studying the vast diversity of microbes we have not yet cultured in the lab.  Certainly with the growing importance of the human microbiome in medical studies and the growing application of molecular systematics (e.g., rRNA surveys) to all sorts of aspects of microbiology, the time is ripe for an award in this area.

And who would get an award if one was given.  Well, certainly one of the people should be Carl Woese, who pioneered the use of comparative analysis of the sequences of rRNA genes to the study of systematics of microbes.  Woese of course was responsible for proposing the existence of a third branch in the tree of life - the archaea.  And even if you do not personally believe that the "three domain" tree of life is perfectly correct, Woese and colleagues (e.g., George Fox, who was a coauthor on some of the pioneering papers) were responsible for making microbial systematics a much more rigorous science than it had been.

And I think a good argument could be made for including Norm Pace in this Nobel as he was the one mostly responsible for pushing the sequencing and analysis of rRNA genes for studying microbes in the environment (though I note, others like Mitch Sogin also helped pioneer this field).  There is a direct path from Woese through Pace to much of modern molecular studies of microbes in the environment, including the latest approach - metagenomics.  In fact, there has even been a Nobel Prize already given that depended on much of this work - the one in 2005 to Barry Marshall and Robin Warren for discovery of the role of Helicobacteri pylori in causing stomach ulcers.

Anyway - just a short post about this - maybe more later.  But I sincerely think this would be a well deserved area in which to hand out one of those Nobel Prizes.  Not holding my breath, but always hopeful.

Here some potentially related things that I have written that may be useful to read:

• Review I wrote in PLoS Biology on molecular studies of microbes
• Other posts I have written about this or related topics:
• A complete summary (probably TMI) about my time at the human #microbiome meeting #HMP2010
• Slides/audio of talk I gave including some of the history of this topic
• Most important paper ever in microbiology? Woese & Fox, 1977, discovery of archaea
• Story Behind the Nature Paper on 'A phylogeny driven genomic encyclopedia of bacteria & archaea' #genomics #evolution

#PLoSOne paper keywords revealing: (#Penis #Microbiome #Circumcision #HIV); press release misleading

UPDATE - READ COMMENTS - LEAD AUTHOR HAS GOTTEN PRESS RELEASE CHANGED

A new paper just showed up on PLoS One and it has some serious potential to be important The paper (PLoS ONE: The Effects of Circumcision on the Penis Microbiome) reports on analyses that show differences in the microbiota (which they call the microbiome - basically what bacterial species were present) in men before and after circumcision. And they found some significant differences. It is a nice study of a relatively poorly examined subject - the bacteria found on the penis w/ and w/o circumcision. This is a particularly important topic in light of other studies that have shown that circumcision may provide some protection against HIV infection.

In summary here is what they did - take samples from men before and after circumcision. Isolate DNA. Run PCR amplification reactions to amplify variable regions of rRNA genes from these samples. Then conduct 454 sequencing of these amplified products. And then analyze the sequences to look at the types and #s of different kinds of bacteria.

What they found is basically summarized in their last paragraph

"This study is the first molecular assessment of the bacterial diversity in the male genital mucosa. The observed decrease in anaerobic bacteria after circumcision may be related to the elimination of anoxic microenvironments under the foreskin. Detection of these anaerobic genera in other human infectious and inflammatory pathologies suggests that they may mediate genital mucosal inflammation or co-infections in the uncircumcised state. Hence, the decrease in these anaerobic bacteria after circumcision may complement the loss of the foreskin inner mucosa to reduce the number of activated Langerhans cells near the genital mucosal surface and possibly the risk of HIV acquisition in circumcised men."

And this all sounds interesting and the work seems solid. I note that some friends / colleagues of mine were involved in this including Jacques Ravel who used to be at TIGR and now is at U MD and Paul Kiem who is associated with TGen in Arizona. For anyone interested in HIV, the human microbiome, circumcision, etc, it is probably worth looking at.

However, the press release I just saw from TGen really ticked me off. The title alone did me in "Study suggests why circumcised men are less likely to become infected with HIV". Sure the study did suggest a possible explanation for why circumcised men are less likely to get HIV infections - the paper was justifiably VERY cautious about this inference. They basically state that there are some correlations worth following up.

The press release goes on to say "The study ... could lead to new non-surgical HIV preventative strategies for the estimated 70 percent of men worldwide (more than 2 billion) who, because of religious or cultural beliefs, or logistic or financial barriers, are not likely to become circumcised." Well sure, I guess you could say that. I think they are iplying you could change the microbiome somehow and therefore protect from HIV but that implies (1) that there really is a causal relationship between the microbial differences in HIV protection and (2) that one could change the microbiome easily, which is a big big stretch given how little we know right now.

Anyway - the science seems fine and not over-reaching. But the press release is annoying and misleading. Shocking I know. But this one got to me.

UPDATE - SEE COMMENTS HERE AND IN FRIENDFEED. LEAD AUTHOR GOT PRESS RELEASE CHANGED.



Price, L., Liu, C., Johnson, K., Aziz, M., Lau, M., Bowers, J., Ravel, J., Keim, P., Serwadda, D., Wawer, M., & Gray, R. (2010). The Effects of Circumcision on the Penis Microbiome PLoS ONE, 5 (1) DOI: 10.1371/journal.pone.0008422

My first PLoS One paper .... yay: automated phylogenetic tree based rRNA analysis

Well, I have truly entered the modern world. My first PLoS One paper has just come out. It is entitled "An Automated Phylogenetic Tree-Based Small Subunit rRNA Taxonomy and Alignment Pipeline (STAP)" and well, it describes automated software for analyzing rRNA sequences that are generated as part of microbial diversity studies. The main goal behind this was to keep up with the massive amounts of rRNA sequences we and others could generate in the lab and to develop a tool that would remove the need for "manual" work in analyzing rRNAs.

The work was done primarily by Dongying Wu, a Project Scientist in my lab with assistance from a Amber Hartman, who is a PhD student in my lab. Naomi Ward, who was on the faculty at TIGR and is now at Wyoming, and I helped guide the development and testing of the software.

We first developed this pipeline/software in conjunction with analyzing the rRNA sequences that were part of the Sargasso Sea metagenome and results from the word was in the Venter et al. Sargasso paper. We then used the pipeline and continued to refine it as part of a variety of studies including a paper by Kevin Penn et al on coral associated microbes. Kevin was working as a technician for me and Naomi and is now a PhD student at Scripps Institute of Oceanography. We also had some input from various scientists we were working with on rRNA analyses, especially Jen Hughes Martiny

We made a series of further refinements and worked with people like Saul Kravitz from the Venter Institute and the CAMERA metagenomics database to make sure that the software could be run outside of my lab. And then we finally got around to writing up a paper .... and now it is out.

You can download the software here. The basics of the software are summarized below: (see flow chart too).

• Stage 1: Domain Analysis


• Take a rRNA sequence
• blast it against a database of representative rRNAs from all lines of life
• use the blast results to help choose sequences to use to make a multiple sequence alignment
• infer a phylogenetic tree from the alignment
• assign the sequence to a domain of life (bacteria, archaea, eukaryotes)


• Stage 2: First pass alignment and tree within domain
• take the same rRNA sequence
• blast against a database of rRNAs from within the domain of interest
• use the blast results to help choose sequences for a multiple alignment
• infer a phylogenetic tree from the alignment
• assign the sequence to a taxonomic group


• Stage 3: Second pass alignment and tree within domain
• extract sequences from members of the putative taxonomic group (as well as some others to balance the diversity)
• make a multiple sequence alignment
• infer a phylogenetic tree

From the above path, we end up with an alignment, which is useful for things such as counting number of species in a sample as well as a tree which is useful for determining what types of organisms are in the sample.

I note - the key is that it is completely automated and can be run on a single machine or a cluster and produces comparable results to manual methods. In the long run we plan to connect this to other software and other labs develop to build a metagenomics and microbial diversity workflow that will help in the processing of massive amounts of sequence data for microbial diversity studies.

I should note this work was supported primarily by a National Science Foundation grant to me and Naomi Ward as part of their “Assembling the Tree of Life” Program (Grant No. 0228651). Some final work on the project was funded by the Gordon and Betty Moore Foundation through grant #1660 to Jonathan Eisen and the CAMERA grant to UCSD.

Wu, D., Hartman, A., Ward, N., & Eisen, J. (2008). An Automated Phylogenetic Tree-Based Small Subunit rRNA Taxonomy and Alignment Pipeline (STAP) PLoS ONE, 3 (7) DOI: 10.1371/journal.pone.0002566

Retraction - I was buzzing off about bees before my time

Recently I wrote a post about the recent study on bees associated with colony collapse disorder. After receiving a well thought out email from one of the authors on the study I have decided to retract my blog and apologize to the authors of the bee study. I rushed out my blog without really considering the evidence and the data very carefully and accept that I screwed this one up big time. The study is much more complex and comprehensive that I led people to believe. In part this was due to lack of detail in the actual manuscript but alas most of the fault lies with me - in not trying to contact the authors for more detail before mouthing off.

So I am giving myself a new award - the genomic jerk award. Hopefully there will be no more recipients.