Does phylogeny matter? (In Eco-Evo meta-analyses) ... Apparently, yes, but it depends.

As many may know - I am pretty obsessed with the uses of phylogeny in biological studies.  In fact, one could say this has driven almost all of my work.  Thus when an email went around a little bit ago about an article for a journal club at UC Davis where the title begins with "Does phylogeny matter?", well, I had to take a look.  Alas, I was a bit worried when I saw the article was in Ecology Letters because I am at home and was not sure about access policies for this journal.

But I was pleasantly surprised to get full access without any library - VPN login to the following article: Does phylogeny matter? Assessing the impact of phylogenetic information in ecological meta-analysis - Chamberlain - 2012 - Ecology Letters - Wiley Online Library.  I cannot figure out WHY it is freely available right now, nor how long it will be, but I took the chance to look the article over.

And I was even more pleasantly surprised to look over the article.  Many meta-analyses can seem forced - if not almost unbearable to look through.  But this one is very well done.  Basically they did a massive comparison of conclusions that one could reach when one either does or does not take into account the phylogenetic non-independence of taxa when conducting meta-analyses in evo-eco studies.  They searched published literature for meta-analyses and then .. well I will use their words here (from the end of their introduction):

Bacteria & archaea don't get no respect from interesting but flawed #PLoSBio paper on # of species on the planet

Uggh. Double uggh. No no. My first blog quadruple uggh. There is an interesting new paper in PLoS Biology published today. Entitled "How many Species Are There on Earth and in the Ocean?" PLoS Biol 9(8): e1001127 - it is by Camilo Mora, Derek Tittensor, Sina Adl, Alastair Simpson and Boris Worm. It is accompanied by a commentary by none other than Robert May, one of the greatest Ecologists of all time: PLoS Biology: Why Worry about How Many Species and Their Loss?

I note - I found out about this paper from Carl Zimmer who asked me if I had any comments.  Boy did I.  And Zimmer has a New York Times article today discussing the paper: How Many Species on Earth? It’s Tricky.  Here are my thoughts that I wrote down without seeing Carl's article, which I will look at in a minute.

The new paper takes a novel approach to estimating the number of species. I would summarize it but May does a pretty good job:

"Mora et al. [4] offer an interesting new approach to estimating the total number of distinct eukaryotic species alive on earth today. They begin with an excellent survey of the wide variety of previous estimates, which give a range of different numbers in the broad interval 3 to 100 million species"

....

"Mora et al.'s imaginative new approach begins by looking at the hierarchy of taxonomic categories, from the details of species and genera, through orders and classes, to phyla and kingdoms. They documented the fact that for eukaryotes, the higher taxonomic categories are “much more completely described than lower levels”, which in retrospect is perhaps not surprising. They also show that, within well-known taxonomic groups, the relative numbers of species assigned to phylum, class, order, family, genus, and species follow consistent patterns. If one assumes these predictable patterns also hold for less well-studied groups, the more secure information about phyla and class can be used to estimate the total number of distinct species within a given group."

The approach is novel and shows what appears to be some promise and robustness for certain multicellular eukaryotes. For example, analysis of animals shows a reasonable leveling off for many taxonomic levels:

One of my new favorite things: paleovirology

Just a quick post here about a paper that came out about a month or so ago: PLoS Biology: Genomic Fossils Calibrate the Long-Term Evolution of Hepadnaviruses

This paper, by Clément Gilbert, Cédric Feschotte is quite cool.  In it they describe their work on "Paleovirology" where they look for viruses than have "endogenized" by inserting into the genome of some host species.  This endogenization is important in particular when the endogenous form becomes inactive and thus, in essence, trapped in the genome.  This in turn is important because many viruses evolve so rapidly when they are "free" that it is very hard to reconstruct their ancient history through comparative analysis.  But the endogenized viruses serve in essence as a molecular "fossil record" that aids in the comparison and phylogenetic analysis of various families of viruses.  As we get more and more genomes, this searching for and analysis of endogenous viruses will get much better.

Anyway, in the paper they report on endogenous viruses in the Zebra Finch genome that are in the Hepadnaviridae family.  Here is their summary:

Paleovirology is the study of ancient viruses and the way they have shaped the innate immune system of their hosts over millions of years. One way to reconstruct the deep evolution of viruses is to search for viral sequences “fossilized” at different evolutionary time points in the genome of their hosts. Besides retroviruses, few virus families are known to have deposited molecular relics in their host's genomes. Here we report on the discovery of multiple fragments of viruses belonging to the Hepadnaviridae family (which includes the human hepatitis B viruses) fossilized in the genome of the zebra finch. We show that some of these fragments infiltrated the germline genome of passerine birds more than 19 million years ago, which implies that hepadnaviruses are much older than previously thought. Based on this age, we can infer a long-term avian hepadnavirus substitution rate, which is a 1,000-fold slower than all short-term substitution rates calculated based on extant hepadnavirus sequences. These results call for a reevaluation of the long-term evolution of Hepadnaviridae, and indicate that some exogenous hepadnaviruses may still be circulating today in various passerine birds.

Figure 4. Summary of the evolutionary scenario inferred in this study.

It is an interesting paper and worth a look if for those who have any interest in viral evolution. And I am becoming more and more fascinated by "Paleovirology" these days so I thought I would just post about this article here.  And I guess I am not alone in this opinion that the article is interesting (though I am late).  Here is some coverage of their paper:

• Ancient Virus Found Hiding Out in Finch Genome
• Ancient viruses lurk in songbird's DNA
• Ancient "Fossil" Virus Shows Infection to Be Millions of Years Old
• Fossil virus leaves evolutionary footprints in songbird DNA
• It's all in the genes: Songbirds have fossil viruses in their DNA ...
• Ancient Bird Virus « Life « Science Today: Beyond the Headlines

Gilbert, C., & Feschotte, C. (2010). Genomic Fossils Calibrate the Long-Term Evolution of Hepadnaviruses PLoS Biology, 8 (9) DOI: 10.1371/journal.pbio.1000495

Cool paper, & winner of "worst new omics word award": Predatosome

And the bad new omics words keep streaming in. Today's winner of the "Worst New Omics Word Award" is going to Carey Lambert, Chien-Yi Chang, Michael J. Capeness and R. Elizabeth Sockett from Nottingham for their use/ invention of "Predatosome". They use this term in the title of their new PLoS One paper: The First Bite— Profiling the Predatosome in the Bacterial Pathogen Bdellovibrio. Here is the very long sentence where the define it:

The gene products required for the initial invasive predatory processes have not been extensively studied but the genome sequencing of B. bacteriovorus HD100 [1] revealed a genome of 3.85Mb, including a core genome similar to that of non-predatory bacteria and some 40% of the genome comprising a potential predicted “predatosome” of genes, encoding both hydrolytic products that may be employed in prey degradation, and genes that may be required specifically for host predation and thus are not conserved across the Proteobacteria.

The paper is actually quite interesting. They use genomic approaches to characterize a fascinating organism - the bacterial species Bdellovibrio bacteriovorus. This bug is a predatory organism - eating other bacteria. Since it eats them from the inside out, some, including these authors, refer to this organism as a pathogen of other bacteria and their is some discussion here and elsewhere for its potential to serve as a "living antibiotic" in much the same way people are trying to use bacterial viruses (a.k.a. phage).

The paper overall is quite nice on first read. They used microarray studies to characterize gene expression patterns in different phases of the life cycle (see Figure above for the life cycle outline). They backed up this work by quantitative PCR studies and regular RT PCR. And based upon their analysis they found some genes that are "Up-Regulated in Predatory, but Not HI" phase (HI stands for host-independent). And here is where they really tell us what they mean by predatosome:

This category of 240 genes are very interesting as they potentially exclude those genes simply involved with release from attack-phase into growth, namely they should be part of the “predatosome” of predatorily specific genes.

It seems to me this terminology is completely unnecessary. All they need to do is say they are studying the genes related to the predatory phase. To assign these genes to the "predatosome" is a bit much. They continue in the paper to report some really interesting stuff. For example, they also examine another predatory bacterial species, and look at whether there are genes conserved in the process between species. They made some really nice figures by the way about the different phases of hte life cycle in this organism and which genes are expressed:

Anyway - the science in the paper is nice. However, the invention of yet another omics word is a bit much. And thus Lambert et al. are winners of the highly coveted "Worst New Omics Word Award" for their invention of "predatosome". Details on the paper are below - and that is where the figures come from too. (Hat tip to Bora for letting me know about the paper, and the word).

Lambert, C., Chang, C., Capeness, M., & Sockett, R. (2010). The First Bite— Profiling the Predatosome in the Bacterial Pathogen Bdellovibrio PLoS ONE, 5 (1) DOI: 10.1371/journal.pone.0008599

Friendfeed comments below:

Confronting Intelligent Design arguments directly in the scientific literature

A representative from Wiley publishing sent me a link to an interesting new paper. Entitled "Using Protistan Examples to Dispel the Myths of Intelligent Design" by Mark Farmer, from the University of Georgia and Andrea Habura, from the University at Albany, New York. It is from the Journal of Eukaryotic Microbiology and is based upon a presentation they gave at a workshop at a conference.

Basically, the article is a detailed discussion of how examples relating to microbial eukaryotes (I hate the term protist ...) that are used by Intelligent Design advocates are, well, BS. And the article discusses the evidence that refutes the ID arguments.

One thing they discuss is the issue of the Cambrian Explosion. ID supporters, such as Stephen Meyer have made many arguments about they feel the diversification in the Cambrian is not explainable through evolutionary processes. Farmer and Habura refute this by pointing out that the diversity seen in microbial eukaryotes at the time of the Cambrian was immense and that what came out of the "explosion" was actually not that spectacular relative to what already existed in the microbial eukaryotes:

The extant diversity of the protists should therefore be seen as the "background radiation" of the eukaryotic Big Bang, with the Cambrian radiation of the metazoa being a subsequent event within a specific group.

They go on to discuss examples involving speciation, the fossil record, evolution of drug resistance in Plasmodium, and a few other things. In each case they discuss a claim by ID supporters and then discuss evidence for why this claim is not valid. Overall the paper is worth reading if you are involved in any discussions with ID supporters.


I note that when I finished the above writing, I went to look at Pubmed to find other examples of people taking on ID arguments in the literature with a focus on issues in microbes. Here are two other recent examples:

• Introduction: Protistan Biology, Horizontal Gene Transfer, and Common Descent Uncover Faulty Logic in Intelligent Design.
• Bacterial flagella and Type III secretion: case studies in the evolution of complexity.

Some discussion of this has now popped up on the web:

• Protozoans Against Intelligent Design
• Genome Web "This might count as preaching to the choir"

FARMER, M., & HABURA, A. (2010). Using Protistan Examples to Dispel the Myths of Intelligent Design Journal of Eukaryotic Microbiology, 57 (1), 3-10 DOI: 10.1111/j.1550-7408.2009.00460.x

#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

World's Smallest Genome of a Cellular Organism?

A one page paper in Science reports on what I think is one of the most exciting findings in microbial genomics in years. The reports describes the sequencing and analysis of the genome of a bacterial endosymbiont of an aphid. This bacteria, known as Carsonella, has a TINY genome - only 160 kbp in length. This is ~ 3 fold smaller than the previously known smallest genome - that of Nanoarchaeum equitans which has a genome of 490 kbp.

I think almost certainly this symbiont should be considered an organelle. It is missing many cellular functions found even in the most reduced symbionts. Thus in essence it may not be the smallest genome of a cellular organism. But who cares how we define it. If it is a new organelle - that is amazing. If it is a tiny cellular genome - that is amazing too.

One thing that strikes me as strange is the fact that the paper is only one page long. It contains so little detail on what was done and what was found in the genome that the story is woefully incomplete. This I would guess is somehow related to a rush to publish but also likely due to it being published in Science, which has severe page restrictions.

This paper has been getting ENORMOUS press coverage for valid reasons. But I agree with Craig Venter (see the New Scientist article) that this genome is not of much relevance to efforts to create a "minimal" genome. This is because the ideal minimal genome is one that can support independent life. Carsonella, is far from independent and thus represents a really wild evolutionary story, but nothing of much relevance to minimal genome studies.

Some related links:

• Commentary by Siv Andersson
• NPR Story
• Nancy Moran's Home Page (one of the corresponding authors of the paper)
• World Science article

Nakabachi, A., Yamashita, A., Toh, H., Ishikawa, H., Dunbar, H., Moran, N., & Hattori, M. (2006). The 160-Kilobase Genome of the Bacterial Endosymbiont Carsonella Science, 314 (5797), 267-267 DOI: 10.1126/science.1134196

Tackling the hairy beast - Tetrahymena genome

Just thought I would put out a little self-promotional posting here on a paper we have published today on the genome of a very interesting organism called Tetrahymena thermophila. This organism is a single-celled eukaryote that lives in fresh water ponds.

This species has served as a powerful model organism for studies of the workings of eukaryotic cells. Studies of this species have led to some fundamental discoveries about how life works. For example, telomerase, the enzyme that helps keep the ends of linear chromsomes from degrading, was discovered in this species. This may not seem too important, but many folks think that degradation of chromosome ends in humans is involved in aging. Perhaps even more importantly, (to me at least) studies of this species were fundamental to the discovery that RNA can be an enzyme. This discovery of catalytic RNA revolutionized our understanding of how cells work and how life evolved. Tom Cech and Sidney Altman were given the Nobel Prize in 1989 for this discovery.

Many (including myself) believe that having the genome sequence of this species will further spur research and its use as a model organism. In addition, we believe that some of the findings we report in our paper will further cement the importace of this species. For example, this species, though single celed, encodes nearly as many proteins as humans and possesses many processes and pathways shared with animals but missing from other model single celled species.

The project that led to this publication was undertaken while I was at TIGR (The Institute for Genomic Research) and involved a collaboration among people at dozens of research institutions around the world. It all started in 2001 when Ed Orias and his colleagues sought to see if anyone at TIGR would be interested in putting in a grant to sequence this species' genome. I responded to the email saying I was interested, especially since I had interacted with multiple people who used this species as a model system (e.g., Laura Landweber at Princeton and Laura Katz at Smith). So I went to a FASEB meeting where the Tetrahymena Genome Steering Committee was meeting and discussed with them how TIGR might help sequence the genome. And after talking to other genome centers, they selected TIGR to put in a grant proposal with them.

We ended up getting funding from two grant proposals - one from NIGMS and the other from the NSF Microbial Genome Sequencing Program. The sequencing was done in a rapid burst at the new Joint Technology Center which TIGR shares with the Venter Institute. And then we spent ~1.5 years analyzing the sequence data (and assemblies) that came out and in the end we fortunately were able to get our paper into PLoS Biology, in my opinion the best place available to publish biology research.

Importantly PLoS Biology is Open Access which allows anyone anywhere to read about our work. This goes well with the free and open release we made of the genome sequence data. In fact, many people published papers on the genome before we did (sometimes scooping us). In the end, I accepted the risks of releasing the genome data with no restrictions inexchange for advancing research on this organisms. I think this risk was well worth it as we still got our big paper published and the field has advanced more rapidly than if we had not released the data.

Other links that may be of interest to people:

• The Tetrahymena Genome Database
• Tetrahymena Web Links
• Ed Orias' Tetrahymena Site
• Tetrahymena site at TIGR

Eisen, J., Coyne, R., Wu, M., Wu, D., Thiagarajan, M., Wortman, J., Badger, J., Ren, Q., Amedeo, P., Jones, K., Tallon, L., Delcher, A., Salzberg, S., Silva, J., Haas, B., Majoros, W., Farzad, M., Carlton, J., Smith, R., Garg, J., Pearlman, R., Karrer, K., Sun, L., Manning, G., Elde, N., Turkewitz, A., Asai, D., Wilkes, D., Wang, Y., Cai, H., Collins, K., Stewart, B., Lee, S., Wilamowska, K., Weinberg, Z., Ruzzo, W., Wloga, D., Gaertig, J., Frankel, J., Tsao, C., Gorovsky, M., Keeling, P., Waller, R., Patron, N., Cherry, J., Stover, N., Krieger, C., del Toro, C., Ryder, H., Williamson, S., Barbeau, R., Hamilton, E., & Orias, E. (2006). Macronuclear Genome Sequence of the Ciliate Tetrahymena thermophila, a Model Eukaryote PLoS Biology, 4 (9) DOI: 10.1371/journal.pbio.0040286

Glassy winged sharpshooter symbionts

For those interested in wine production, or symbioses, you maybe interested in a paper we published a few days ago. It was on a study we did of bacterial symbionts of an insect known as the glassy winged sharpshooter. This insect is a vector for Pierce's Disease in grapes - a nasty disease that if it is found in a vineyard might lead to the vineyard being sacrificed for the greater good.

Anyway - we did a study of bacteria that live inside the insect that make nutrients for their insect host and without whcih the insect will die. An understanding of these symbionts will hopefully lead to better methods to control the spread of this invasive insect.

Our paper can be found in PLoS Biology here.
A synopsis of our article is here.
An article in Science Now about our study is here
An article in the Central Valley Business Times is here.
Nature highlighted it in their "Research highlights" section
And ASM article about this here
Link to our collaborator's lab (Nancy Moran)

Some new links about our paper

• SF Sourcebook
• Complex Medium
• Sacramento News and Review article
  

For more information about the sharpshooter and Pierce's Disease seee the following links

• Pierce's Disease Control Program for State of California: here
• Glassy winged sharpshooter media information here
• Introduction to Pierce's Disease here

Wu, D., Daugherty, S., Van Aken, S., Pai, G., Watkins, K., Khouri, H., Tallon, L., Zaborsky, J., Dunbar, H., Tran, P., Moran, N., & Eisen, J. (2006). Metabolic Complementarity and Genomics of the Dual Bacterial Symbiosis of Sharpshooters PLoS Biology, 4 (6) DOI: 10.1371/journal.pbio.0040188