Posts in category Bioinformatics

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NASA and JCVI host symposium on the evolution of Earth and Life

On May 12th and 13th, the J. Craig Venter Institute in San Diego will be hosting a NASA Astrobiology Institute-funded symposium titled “Paleobiology in the genomics era.” Paleobiology is the study of the origins and evolution of life and, by nature, is interdisciplinary. The goal is to bring together scientists united by this common interest but differentiated by expertise.  A major intellectual challenge to paleobiology is the close interaction between environment and life.  As life evolved, it changed the environment and suffered the consequences.  One of the most extreme examples is the invention of oxygenic photosynthesis by blue-green algae cyanobacteria; the sunlight-fueled production of dramatically changed the availability of crucial elements of life, like nitrogen, sulfur, iron, zinc, copper, and other trace metals.  Genome-based analyses showed that these environmental changes modulated the emergence of metal-requiring proteins.  For example, proteins that bind Fe evolved when the earth was Fe rich. Essentially, one biological event changed the environment, which in turn induced a subsequent biological change; a feedback cycle between biota and planet.

In order to study these interactions in a robust fashion, numerous lines of evidence must be integrated, despite originating from disparate fields like organic and inorganic geochemistry (oils and metals in rocks), micropaleontology (tiny fossils), and evolutionary biology.  Recent years have observed the emergence and maturation of synthetic biology and computational biology, two fields with tremendous potential for the formulation and testing of hypotheses about the evolution of life. To facilitate a dialog between these fields, myself, along with Ariel Anbar from Arizona State University, and John Peters and Eric Boyd from Montana State University, have invited experts to present their work as it pertains to paleobiology.  The topic list almost appears schizophrenic, with numerous hard-core geochemical talks being followed presentations on molecular genetics, synthetic biology, metagenomics, and comparative genomics.   This was intentional. I hope to feel intellectually challenged in the fashion of a 1st year graduate student and further hope that I’m not the only one.  A major wild card at the moment is the identity of over 2/3rd of the attendees.  With travel grants available for graduate students, post-doctoral researchers, and faculty, we hope to incorporate novel perspectives not covered by the confirmed speakers.

While the content of the meeting is exciting, the format is pretty sweet too. As part of NASA’s Workshop Without Walls series, the meeting will be webcast live with an accompanying live stream chat.  Thus, people will be able to see the presentations and pose questions and comments during the attendant discussions.  Previous workshops have often had hundreds of live viewers throughout the meeting, despite only dozens of in situ attendees.   The actual energy savings for a single meeting are modest in isolation; imagine 250 people not flying 500 miles and you basically have a single 737 flight that remains grounded.  However, the future of environmentally-friendly science requires important preliminary steps to change dominant trends.  Similarly, the talks will be streamed live without charge and deposited in the open access scientific podcast site, Scivee.tv; economic barriers to information exchange are removed.

Needless to say, I’m looking forward to this meeting. Organizing something like this is an absolute undertaking. The number of details that need attention is astounding.  And if you think I actually could do that, you don’t know me.  Numerous people at JCVI have provided invaluable assistance, including Matt LaPointe and Jasmine Pollard, Robert Friedman, Dave Negrotto, and Jody Wilson.  It would also have no chance of happening if it not for Pat Goley, who has handed the numerous (re: uncountable) details I’ve lapsed on.

Check out the NASA page for the meeting and webcast registration.

http://astrobiology.nasa.gov/nai/geobiology2011

JCVI Supports Human Mircrobiome Body Site Experts with Shotgun Data Analysis

Members of the Human Microbiome Project (HMP) Consortium (see http://commonfund.nih.gov/hmp and http://www.hmpdacc.org for more information on the project and partners) including human microbiome body site experts gathered for a virtual Jamboree January 19th. The fully online-based Jamboree has been set-up to communicate initial data products and tools best suited for analysis, primarily to make the data amendable/consumable in a user-friendly way for body site exerts. 61 participants followed the Jamboree agenda with presenters given access to a common desktop that was shared via the internet using an online collaboration tool. Results from  the Data Analysis Working Group (DAWG) were presented in the areas of 16S rRNA gene sequence (16S DAWG) and metagenomic whole-genome shotgun analysis (WGS DAWG). The efforts of the 16S DAWG focus on marker-gene based approaches to estimate biological diversity and how marker variability is associated with patient meta-data. The WGS DAWG  complements results from the 16S marker based analysis with comprehensive sequencing of random pieces of genomic DNA from the collection of microorganisms which inhabit a particular site on, or in, the human body (microbiome). These analyses allow researchers to investigate among other questions what microorganisms are present, and the nature and extent of their collective metabolism, at a particular body site. Ultimately researchers want to relate this information to healthy versus diseases states in humans.

METAREP tutorial presented as part of the HMP Virtual Jamboree

The current survey comprises more than 700 samples from hundreds of individuals taken from up to 16 distinct body sites. Illumina sequencing has yielded more than 20 billion Illumina reads and annotation data produced from the sequences exceeds 10 terabytes. In anticipation of such data volumes, we have developed JCVI Metagenomics Reports (METAREP), an open source tool for high-performance comparative analysis, in 2010. The tool enables users to slice and dice data using a combination of taxonomic and functional/pathway signatures. To demonstrate how the tool can be used by body site experts, we picked and loaded sample data from 17 oral samples and presented a quick tutorial on how users can view, search, browse individual samples and compare multiple samples (see video). The functionality was very well received and body site experts asked JCVI to make all the 700+ samples available. As a result of the Jamboree, JCVI in agreement/collaboration with the HMP Data Analysis and Coordination Center and the rest of the HMP consortium, will soon set-up a dedicated HMP METAREP instance that will allow body-site experts and eventually other users to analyze the DAWG data in a user-friendly way via the web.

A Look Back at 2010 at the JCVI…

As the J. Craig Venter Institute (JCVI) soars into its 19th year, we reflect on the past year of highlights and accomplishments to mark the close 2010 and look forward to more significant scientific advances in 2011.

JCVI Top 10 of 2010 …

1. First Synthetic Cell: Fifteen years in the making, 2010 brought to bear with huge anticipation the successful construction of the first self-replicating, synthetic bacterial cell. The work was published in Science in May. The synthetic cell called Mycoplasma mycoides JCVI-syn1.0 is the proof of principle that genomes can be designed in the computer, chemically made in the laboratory and transplanted into a recipient cell to produce a new self-replicating cell controlled only by an artificial genome. Although the first synthetic cell was not designed to produce a specific bioproduct, the team has shown that this can be done and the potential benefits are numerous. The research team, lead by JCVI President Craig Venter, Hamilton Smith, Clyde Hutchison, and Daniel Gibson, envision a future where the rapid design and production of biological products using synthetic biology techniques will be used to produce clean fuels, medicines, and other bioproducts. Throughout the course of this work, the JCVI Policy group has extensively engaged in outside review of the ethical and societal implications of this work, including advising the new Presidential Commission on Bioethics on their recommendations for oversight.

M. mycoides JCVI-syn1

M. mycoides JCVI-syn1

2. Synthetic Vaccines: Following on the heels of the announcement of the first synthetic cell, the company Synthetic Genomics Inc. and JCVI announced in October the formation of a new company, Synthetic Genomics Vaccines Inc. (SGVI). The privately held company is focused on developing next generation vaccines that can be rapidly produced and tested, which is especially important for outbreaks of new infectious diseases. SGVI also announced a three-year collaboration with Novartis to apply synthetic genomics technologies to accelerate the production of the influenza (flu) seed strains required for vaccine manufacturing. The seed strain is the starter culture of a virus, and is the base from which larger quantities of the vaccine virus can be grown. Under this collaboration, Novartis and SGVI will work to develop a “bank” of synthetically constructed seed viruses ready to go into production as soon as WHO makes recommendations on the flu strains. The technology could reduce vaccine production time by up to two months, which is particularly critical in the event of a pandemic.

3. Hydra Genome – one of the animal kingdom’s earliest common ancestors: JCVI scientists along with more than 70 other researchers from around the world, have sequenced and analyzed the genome of Hydra magnipapillata, a fresh water member of the cnidaria– stinging animals that include jellyfish, sea anemones and corals. The research, published in the March 14 edition of Nature, was co-led by Ewen F. Kirkness, JCVI, Jarrod A. Chapman, Department of Energy Joint Genome Institute, and Oleg Simakov, University of California, Berkeley. This is the second sequenced cnidarian genome, following that of a sea anemone, Nematostella vectensis, in 2007. The ancestors of these two species diverged more than 500 million years ago, and comparison of their genomes has revealed common features of the earliest animals that gave rise to the diversity of animals on Earth today. The team found clear evidence for conserved genome structure between the Hydra and other animals, like humans. Unexpectedly, the sequencing also revealed a novel bacterium that lives in close association with the Hydra.

4. Uncovering the Human Microbiome: Microbes are living within and on the human body and this collective community is called the human microbiome. JCVI Scientists, as one component of the large scale NIH Roadmap Human Microbiome Project, and along with colleagues at three other genome centers sequenced the genomes of ~180 microbes from the human body, published in the May 21 edition of Science. At the JCVI we anticipate sequencing an additional 400 species over the next few months. Colleagues at the JCVI are also using single cell approaches to isolate new strains that have not been cultured – isolates whose genomes will also be completely sequenced. The role these microbes play in human health and disease is still relatively unknown and these approaches are allowing us to gain a greater understanding of these enigmatic species.

5. Body Louse Genome: A global research team led by Ewen Kirkness and colleagues from JCVI published a study in the Proceedings of the National Academy of Sciences in June describing the sequencing and analysis of the human body louse, Pediculus humanus humanus, a human parasite responsible for the transmission of bacteria that cause epidemic typhus, relapsing fever and trench fever. Detailed analysis of the genome was then conducted by a large international group of 71 scientists, coordinated by Barry Pittendrigh, University of Illinois, and Professor Evgeny Zdobnov, University of Geneva Medical School. Comparative studies of the body louse genome with other species revealed features that will enhance our understanding of the relationships between disease-vector insects, the pathogens they transmit, and the human hosts. In addition to the targeted louse genome, the project unexpectedly yielded the complete genome sequence of a bacterial species, Riesia, that lives in close association with lice, and which is essential for survival of the insects. The researchers believe that the genome will be a valuable reference for evolutionary studies of insect species, especially in the areas related to insect growth and development.

6. Castor Bean Genome Sequencing: A research team co-led by Agnes P. Chan and colleagues from JCVI and Jonathan Crabtree and others at the Institute for Genome Sciences, University of Maryland School of Medicine, published the sequence and analysis of the castor bean (Ricinus communis) genome in Nature Biotechnology in August. Because of the potential use of castor bean as a biofuel and its production of the potent toxin ricin, the team focused efforts on analysis of genes related to oil and ricin production. The analyses could be important for comparative studies with other oilseed crops, and could also allow for genetic engineering of castor bean to produce oil without ricin. Identifying and understanding the ricin–producing gene family in castor bean will be important in preventing and dealing with potential bioterrorism events. Genomics enables enhanced diagnostic and forensic methods for the detection of ricin and precise identification of strains and geographical origins. As a next step, the group suggests further comparative genomic studies with the close relative cassava, a major crop in the developing world, to further elucidate their disease resistance aspects.

7. Science Education: JCVI was an Official Partner of the inaugural USA Science and Engineering Festival held on the National Mall in Washington, DC in October. The Festival, which was the country’s first national science festival, included over 500 of the country’s leading science and engineering organizations with the aim to reignite the interest of our nation’s youth in the sciences. The JCVI ‘Discover Genomes’ Bus was showcased during a two-day expo and some of the research being done at JCVI was presented to around 1700 visitors by our scientists and staff.

There were lines all day!

8. Viral Genomics– In 2010 the JCVI has published over 1600 influenza genomes and over 75% of all published flu genomes to date have been sequenced by the JCVI, totaling over 6000 genomes. This year the diversity of viral genomes we have sequenced has significantly expanded under the NIH Genomic Sequencing Center for Infectious Diseases contract. Some of the projects include viruses causing diseases such as measles, mumps, rubella, encephalitis, SARS, and the common cold, just to name a few. The viral group has annotated and published 79 Rotavirus (stomach flu) and 33 Coronavirus genomes (includes SARS and common cold) this year and many more will be published in 2011. The pace of sequencing and finishing genomes has also increased this year as a result of adoption of nextgen platforms (e.g. Illumina/454 and Illumina/Solexa) and the development of more efficient methodologies to increase productivity while reducing costs.

9. Marine Microbial Genome Sequencing Project: JCVI scientists have continued their quest to isolate and sequencing microbes living in global ocean waters to discover new genes and enzymes, and to help understand the role microbes play in the ocean ecosystem. Shibu Yooseph, Kenneth Nealson and colleagues at JCVI published an analysis of 137 known marine microbial genomes living in the global ocean surface in Nature in November. These genomes were compared to metagenomic samples of ocean waters of 10.97 million sequences of JCVI’s Sorcerer II Global Ocean Sampling (GOS) metagenomic data and thousands of 16S rRNA sequences. The marine genomes were collected as part of the Gordon and Betty Moore Foundation-funded Marine Microbial Genome Sequencing Project, a project coordinated by JCVI that has a primary goal of obtaining whole genome sequences of ecologically important microbes from a variety of diverse, global marine environments. The work provides a good example of combining metagenomic data with sequenced genomes data to study microbial communities and to generate testable hypotheses in microbial ecology.

10. Sorcerer II Global Ocean Sampling Expedition: On December 17th 2010 Sorcerer II arrived in Florida after spending the last two years with her crew collecting samples in The Baltic, Mediterranean and Black Seas. Funded generously by the Beyster Family Foundation Fund, The San Diego Foundation, and Life Technologies Foundation, Sorcerer II has sailed ~28,000 nautical miles since departing San Diego in March 2009. During this time 212 samples were collected and over 5,100 liters of sea water was filtered and sent to JCVI for analysis of the microbial life contained within these samples. The JCVI established strong collaborations with scientists in all 16 countries in which samples were collected, which will lead to joint publications and future collaborative studies in the new year. Read more.

Sunrise in the Ligurian Sea

Looking Forward to 2011…

Ten-year anniversary of the Human Genome Project: To commemorate the anniversary of the publications of the first human genome sequences in 2001, JCVI and Nature are hosting a conference and celebration in February 2011 titled – Human Genomics: The Next 10 Years. The conference will look forward to the promises of human genomics for the next 10 years, with sessions on medical advances related to genomics; the technological and ethical challenges of human genomics; personalized and familial genomics; the human microbiome project; variation in the human genome; and making sense of the genetic code. This conference will be a great way to jump into the new year and inspire the grandiose ideas and achievements that genomic scientists will accomplish over the years to come.

Lucene Revolution Conference 2010

I arrived late in Boston after my plane from Washington DC was delayed. On the agenda – the next four days the Lucene Revolution conference and a Solr application development workshop organized by Lucid Imagination. The conference promised a unique venue (the first of its kind in the US) to meet developers that all share the same challenge: to enable users to find relevant information in growing bodies of data quickly and intuitively. I was looking forward to hearing many interesting talks given by experts of the field, to learning how to build intuitive search interfaces, and to get an idea where things are heading in the next years. As the developer of JCVI’s Metagenomics Reports (METAREP), I was especially looking forward to the Solr workshop to learn some of the tricks from the experts to tweak the search engine behind this open-source metagenomics analysis tool.

The Early Revolution

But before the revolution could happen and I could enjoy some splendid time at the Washington Dulles airport, Doug Cutting had to start developing a Java based full-text search engine called Lucene in 1997. Lucene became an open-source project in 2000 and an Apache Software Foundation project one year later. In 2004, Solr emerged as an internal CNET project created by Yonik Seeley to serve Lucene powered search results to the company’s website. It was donated by CNET to the Apache Software Foundation in 2006.

Google Trend for Solr

Google Trend for Solr

Early this year, both projects merged and development since then has been carried out jointly under the umbrella of the Apache Software Foundation. Meanwhile many companies use Solr/Lucene, among them IBM, LinkedIn, Twitter, and Netflex. How did this happen?

The Lucid Imagination Solr Application Development Workshop

In search of an answer, I made my way from my hotel to the conference venue, the Hyatt Hotel located along the beautiful Boston harbor bay. The 2-day workshop was a brute-de-force tour of Solr features, configuration, and optimization. It also touched on the mathematical theory behind Lucene’s search result scoring and on evaluating result relevance.  The 2-day workshop covered enough material to warrant a third day. Given this optimistic agenda, there was not much time for the labs (exercises) and the trainer had to focus more on breadth than on depth. As a one-year Solr user, many of the general concepts were familiar so I was more interested in details. A comprehensive hand-book and an excellent exercise compilation came to the rescue and provided me with the needed detail to follow up on subjects that were touched on. There were two parallel Solr classes. In my class, 25 participants followed the training. The mix included developers working for media, defense, and other co-operations. Academia was represented by several libraries and universities.

Solr Application Development Workshop

A powerful feature I had not heard before is the DISMAX RequestHandler. The handler allows to abstract complex queries. Users can enter a simple query without complex syntax or specifying a search field and behind the scenes the handler will do its magic.  It searches across a set of specified fields which (among other things) can be weighted by importance. Additional information about this handler and other snippets I collected during the class can be found in my Solr workshop notes .

The Lucene Revolution Conference

After a mediocre coffee brewed in my hotel room, I headed to the conference venue on the second floor of the Hyatt Hotel. The first day of the conference started with a podium discussion about Cutting Edge of Search that included Michael Busch (Twitter), John Wang (LinkedIn), Joshua Tuberville (eHarmony), and Bill Press (Salesforce.com). The discussion went back and forth showcasing each search platform and the experience in developing it. When asked what he would do differently in retrospect John Wang from LinkedIn ironically mentioned that he would “ban recruiters” – if I correctly remember he mentioned that they “spam-up” the system.

Lightning Talk “Using Solr/Lucene for High-Performance Comparative Metagenomics”

Joshua Tuberville from eHarmony provided valuable advice to developers: “Avoid pet queries for benchmarking a system – use a random set of queries instead.” He also suggested tracking queries that web site users enter for optimization, adding “it surprises me every day that the world is not made up from engineers, but it is a fact.” Avoid unnecessary complexity and duplicating efforts. Use open-source if available. For example, instead of implementing their own Lucene wrapper, eHarmony made use of the open-source project Solr. Bill Press added “Do not be afraid to tear things down, rebuild it many times if needed.”

“Companies do not have time to debug code.” Eric Gries (CEO Lucid Imagination)

Eric Gries, CEO of Lucid Imagination, presented ‘The Search Revolution: How Lucene & Solr Are Changing the World’. In the introduction, he pointed out that Solr/Lucene is the 10th largest community project and the 5th largest Apache Software Foundation project. “Open-source projects need a commercial entity behind them to help them grow”. “Companies need no errors, they do not have time to debug.” His main part focused on his company’s LucidWorks Enterprise software which is based on the open-source project Solr/Lucene. Features that separate it from the open-source version include smart defaults, additional data sources, a REST API that allows programmatic access via Perl/Python/PHP code, standardized error messages, and click based relevance boosting. Later, Brian Pinkerton, also from Lucid Imagination presented additional details. He revealed that their software is based on elements of the upcoming Solr 4.0 version and is fully cloud enabled (added SolrCloud patch). It uses ZooKeeper to manage node configuration and failover. All website communication is done in JSON .  The enterprise version supports field collapsing for distributed search.

“A picture communicates a thousand words but a video communicates a thousand pictures.” Satish Gannu (Cisco)

Satish Gannu from Cisco stressed the increasing prevalence of video data and how such data is changing the world. More and more video enabled devices are pushed on the market. Collaboration is increasingly done across the world. Meetings are recorded and shared globally. Videos are replacing manuals. Cooperate communication/PR via video is increasing. He related the popularity of video to the fact that “A picture communicates a thousand words, but a video communicates a thousand pictures” and that “60% of human communication is non-verbal.” Satish went on to highlight Cicso’s video solutions that make use of automatic voice and face recognition software to store metadata about speakers to enrich the user experience. For example, users can filter out certain speakers when watching recorded meetings. More can be found here.

View of Boston

“Mobile application development will be the driver of open-source innovation.” Bill McQuaide (Black Duck Software)

One of the highlights that morning was Bill McQuaide’s talk on open source trends. Based on diverse sources, including his company Black Duck Software, he showed that software IT spending is down, that 22% of software is open source, and that 40% of software projects use open source. There is an enormous amount of new open source projects targeting the cloud with a lot of competition. Among top open-source licenses are the GNU General Public Licenses, GPL 3.0, and BSD licenses. The three predominant programming languages used by open-source developers are C, C++ and Java. Mobile development will be the driver of innovation in the open-source community especially developments around Google’s Android operating system.  To manage licenses for projects that integrate dozens of open-source projects such as Android and to ship the bundled software to customers can become very complex. For this and other reasons, McQuaide recommends companies and institutions to have policies for implementing open source, integrating third party tools, and identifying and cataloging all open source software used.

Distributed Solr/Lucene using Hadoop

An excellent talk was presented by Rod Cope, from Open Logic.  He presented Real-Time Searching of Big Data with Solr and Hadoop.  The search infrastructure centered around Hadoop’s distributed file system on top of which they cleverly arranged several other technologies.  For example, Hadoop’s HBase database provides fast database lookups but does not provide the power of Lucene text searches. Solr/Lucene however is not as optimized to return stored document information. Their solution is to use Solr/Lucene to search indexed text fields, storing and returning only the document ID.  The returned document ID is then used to fetch additional information from the HBase database. Open Logic uses the open-source software katta to integrate Lucene indices with Hadoop and increased fault tolerance by replicating Solr cores across different machines. Also, corresponding master and slave servers were set up to run on different machines for indexing and searching respectively.  The set up he described runs completely on commodity hardware and new machines can be added on the fly to scale out horizontally.

“It surprises me every day that the world is not made up from engineers but it is a fact.” Joshua Tuberville (eHarmony)

Next on the agenda were seven minute lightning talks. I opened-up the lightning talk session describing our Solr/Lucene based open-source web project METAREP for high-performance comparative genomics (watch). Next was Stefan Olafsson from TwigKit presenting ‘The 7-minute Search UI‘, a presentation which I thought was another gem of this conference. In contrast to other talks, it focused on user experience and intuitive user interfaces. TwigKit has developed a framework that provides well designed search widgets that can be  integrated with several search engines.

“If nobody is against you in open source then you are not right.” Marten Mickos (CEO Eucalyptus)

The key note presentation on the second day was presented by Marten Mickos the CEO of Eucalyptus and former CEO of MySQL. He opened by advocating his philosophy of making money out of open source projects. “Innovation is a change that creates a new dimension in performance” he said and mentioned the open-source Apache web server that allows anyone to run a powerful web server. He added “Market disruption is a change that creates a new level of efficiency” and referred to MySQL originally designed to scale horizontally. While in 1995 such a design was a draw-back compared to other marked solutions, scale-out has become the dominant design today. Now, within the cloud, horizontal scaling is the key. A fact that has made MySQL the most used database in the cloud.

He observed that “while most successful open-source projects are related to building infrastructure software, servers and algorithms, there are only a few open-source projects centered around human behavior, user experience and user interfaces. The latter projects are mainly developed in closed source environments.” Then he went on praising open-source as a driver for innovation “Open source is so effective because you are not protected. Code can be scrutinized by everybody. In a close sourced company, your only competition is within the company, while in open source you compete with everybody.” Open source is a way to innovate and it is more productive. It usually takes a stubborn individual to drive things. Innovation mostly stems from single individuals that are supported by the community.

When asked how to maintain property rights as a company when running an open-source model, he responded “keep things that keep the business going proprietary but open-up others. The key is to be very transparent with your model.”

What’s next ?

In a podium session the core Solr/Lucene committer team discussed future features. The team works on rapid front-end prototyping using the Apache Velocity template engine and Ajax. The prototyping code can be found in the current trunk of the Solr/Lucene code repository under the /browse directory. A Solr/Lucene cloud enabled version is being developed. Twitter’s real time search functionality will be integrated. Other open source projects that are being integrated are Nutch, a web-search software, and Mahout for machine learning (http://mahout.apache.org).  New features will include pivot tables (table matrices), a hierarchical data type, spatial searching, and flexible indexing.

The above represents a subset of talks that took place. There were many other interesting talks – some took place in parallel sessions. Individual presentations can be downloaded from the Lucid Imaginations conference page. A selection of videos is available here. The next Lucene Revolution conference will take place in San Francisco May 2011.

After four days of Solr/Lucene, many coffees, talks, discussions, I left inspired by the conference. It dawned on me that the real revolution is not the search technology but the strong community spirit itself that has emerged and drives developers to jointly work towards a common goal.

Take home message of the 2010 Amebiasis Montreal Meeting: beware of who you kiss…

The Entamoeba community is a small and collegial one.  Everyone knows everyone and everyone else wants to collaborate, and learn and do more to tackle down this neglected among neglected diseases.  For many, the thought of an amoeba brings to memory Garry Larson’s The Far Side amorphous characters watching TV and dealing with domestic issues…but what a few know is that the WHO considers amebiasis one of the major health problems in developing countries surpassed  only by malaria and schistosomiasis for death caused by a parasitic infection.

Amoeba Real Life...

TIGR/JCVI has had a long-standing relationship with Entamoeba histolytica and other related species. Started by Brendan Loftus back in the day, followed by Neil Hall and continuing by yours truly, with the Entamoeba GSCID project, we have provided this eager community with what they are in great need of: genome sequences. And they are appreciative of that, for sure.

The meeting was small and intense, and started with a one day workshop on clinical aspects of the disease, not only the enteric disease, but the oral disease. It was good to mingle with dentists and see their point of view and the devastating reality of clinical cases of periodontal disease…and periodontal disease affects, at one point of another, the majority of the adults. And Entamoeba gingivalis is always there…but not always diagnosed.  Why? We assumed bacteria, bacteria, and bacteria. Treat the bacteria with antibiotics…but ignore the amoebas. Patients do everything right when it comes to oral hygiene, but the disease persists, the bone continues to be destroyed, and teeth are lost…secondary consequences such as fatigue, diabetes, heart disease, renal dysfunction, low birth weight, and a myriad of other diagnoses are known to be related to oral health and periodontal disease, but rarely in the context of this amebiasis infections. Interesting cases of entire families being affected by this parasite were presented (it is highly contagious). Dr. Bonner, the organizer of the conference is a big advocate of microscopes in the dentist practices as the primary diagnostic method for periodontal disease and identification of amebiasis, and he is surely determined to speak to the world about that. And he also wants the genome done.

Entamoeba community, Montreal 2010

During the core of the Meeting, Neil Hall and myself were the only ones on the genomics side of things: we presented on SNP analysis done so far on the strains we have sequenced, in their case using SOLiD, on our case just 454 (for now). Both talks were very well received and the community is eager to see more. Both pieces of work will be used in a global SNP analysis, particularly focusing on a family of proteins known to be involved in virulence, the Gal/GalNac lectins. These proteins are one of the main targets for vaccine development, led by Dr. Bill Petri, who is in the process to start full speed with that endeavor. Also on this topic, Dr. Jonathan Ravdin (Dean and Executive Vice President of the Medical College of Wisconsin) presented the results on an intranasal Gal-lectin subunit vaccine on experimental Entamoeba histolytica in baboons, showing its protective effect against enteric colitis, and the promising future of this vaccine target for humans.

For amebiasis “a la mode”, the usual suspect topics of the conference involved Entamoeba histolyica signaling pathways, classic protein characterization of large families, proteases and invasion, and large genotyping studies of outbreaks, as well as mechanism of pathogenesis.  One interesting talk was on the generation of cyst like forms in vitro for Entamoeba histolytica, a true breakthrough, since there is no model or encystations so far, and the possibility to obtain this structures in vitro will certainly open up a whole new world of studies that before were confined to Entamoeba invadens, a very distant parasite of lizards that does encyst in vitro.

A modest interest into drug discovery was present. There are so far two kind of drug therapies for amebiasis, luminal amoebicides (paromomycin, diloxanide furoate and iodoquinol) for intestinal disease, but ineffective against organisms in tissue and metronidazole and other derivatives,  for invasive disease. However, resistance to these drugs is easily achievable at clinically acceptable drug levels. James McKerrow (UCSF) group presented a high throughput screening of small molecules using an FDA-approved library of drugs and known bioactive compounds, and they identified six compounds with similar or better activity than metronidazole, in vitro. Other group from Mexico is focusing on probiotics and natural compounds such as Astrophitus capricorne (cactus), Jatrhropa dioica and Eucalyptus camaldulensis.

Montreal View from Le Crystal Hotel

To finalize, because the list of interesting things can be just too much, a wonderful piece of work that will hopefully provide an extensive framework for further studies of host susceptibility to E. histolytica comes from Bill Petri’s lab. They have performed a small hairpin RNA (shRNA) screen to identify human factors crucial for E. histolytica cytotoxicity. Using a mammalian shRNA knockdown library they did nine rounds of selection using 1:5 parasite:host and 1:50 parasite:host ratios and resistant clones after 6 rounds of selection, were sequenced using Solexa. This way, they identified a number of host gene families including kinases, surface receptors and ion channels that may be important for susceptibility to the parasite, and of course, they are working on that…

But the take home message that I have imprinted in my brain is…do not kiss your dog.  After three years of age, ALL DOGS have periodontal disease. And inevitably they have Entamoeba gingivalis and possibly other species as well…and they are one of the sources of infection to humans. Dog kisses owner, owner kisses lover, wife, husband, and kids…and the amoeba conquest of the world continues…

For your delight, two movies on the topic: Mark Bonner, the meeting organizer (in French) and a film of a patient with periodontal disease biofilm. Enjoy!

Entamoeba histolytica research presented at the Molecular Parasitology Meeting

Entamoeba histolytica causes invasive intestinal and extraintestinal infections, known as amoebiasis, in about 50 million people and still remains a significant cause of human death in developing countries. However, for unknown reasons, fewer than 10% of E. histolytica infections are symptomatic (causing symptoms such as diarrhea, dysentery or liver abscess). The J. Craig Venter Institute is among the institutions awarded the NIAID Genome Sequencing Centers for Infectious Diseases (GSCID) contracts to provide high-quality genome sequencing and high-throughput genotyping of NIAID Category A-C priority pathogens.

Photo of Entamoeba histolytica

Entamoeba histolytica in the trophozoite stage.

A GSCID project led at JCVI by Dr. Elisabet Caler includes performing whole-genome sequencing of Entamoeba phenotypic variants from symptomatic, asymptomatic and liver abscess-causing strains chosen to include a range of clinical manifestations and taken from human cases, as well as strains grown under different conditions. Our objective is to develop a genome-wide landscape of Entamoeba diversity to understand how sequence variations in the parasite relate to pathogenicity (ability to cause disease) and clinical outcome.

The Molecular Parasitology Meeting held at the Woods Hole Oceanographic Institution, Woods Hole, MA last week provided a window into the exciting science of Parasitology.  The keynote speaker, Fotis Kafatos, spoke on “Major Challenges to Global Health in the Tropics and Beyond–Insect Vectors of Malaria and Other Parasitic or Viral Diseases.”  Dr. Kafatos stressed that a multi-pronged approach to the control of malaria is necessary to prevent the devastating loss of life that malaria causes.

Woods Hole Oceanographic Institution

A view of Woods Hole Oceanographic Institution.

The many excellent papers and posters provided an overview of the field, including   Plasmodium falciparum, Toxoplasma gondii, the trypanosomes, Giardia lamblia, Trichomonas vaginalis, Entamoeba histolytica, Schistosoma species, Babesia bovis, and associated vectors.  Topics spanned basic biology, drug design, sequencing and host-pathogen interactions.

I presented an overview of the Entamoeba sequencing project at the meeting.   Discussions as a result of the presentation included questions about the details of sequencing and handling the next-generation sequencing data.   We had animated discussions about methods for assembly of the DNA sequences, including reference-guided vs de novo assembly.   Many attendees were impressed with JCVI’s open-source METAREP metagenomic tool (J. Goll, et al., Bioinformatics 2010).  Determination of the best methods for the analysis of differences in the clinical isolates generated much discussion.  Entamoeba researchers see the sequences as a great resource and are looking forward to being able to mine the data.  One, from India, was very excited that he was going to have about 15 times the resources he has had in the past, since he has had only had one genome to mine up until now.

The Molecular Parasitology Meeting was an excellent venue for scientific exchange.  The Entamoeba histolytica GSCID project will help us understand the pathogenicity of Entamoeba histolytica, and has the potential to save lives in developing countries.

Virtual Comparative Metagenomics

We have created an open virtualization format (OVF)  package of JCVI’s Metagenomics Reports (METAREP)– a high performance comparative metagenomics analysis tool. The software runs on a web server, retrieves data from two different database systems and uses R for statistical analysis. The new OVF package bundles all these 3rd party tools and is configured to run out of the box in a virtual machine.

Screenshot of the virtual box appliance import wizard. The wizard allows you to specify the CPU and memory usage of the virtual machine on which METAREP will run on.

To run a virtual version of METAREP on your machine, follow these steps

  1. download the METAREP OVF package from our ftp site [download] .
  2. unzipp the OVF package
  3. download and install Oracle’s Virtual Box, a OVF compatible virtualization software [download]
  4. Start Virtual Box
  5. Click File/Import Appliance and select the OVF file.
  6. Adjust RAM/CPU usage using the Appliance Import Wizard (see image)
  7. Start VM
  8. Double-Click on the METAREP firefox link on the VM desktop
  9. Log into METAREP with username=admin and password=admin

This virtual machine appliance is the first step in developing a fully cloud-enabled analysis platform where users can easily launch the application wherever is most convenient: on their personal desktop or in the cloud where they can scale-out the appliance to suite their needs.

Future virtual machine images will be certified to run on other virtualization software platforms. Stay tuned.

If you like to learn more about METAREP and talk to the developers,  join us  at  Lucene Revolution Conference in Boston (October 7-8  2010). We will present a lightning talk about METAREP  the first day of the conference 5pm  (see agenda).

Links:

JCVI’s METAREP Instance

METAREP Flyer

METAREP Manual

METAREP Source Code

HMP Consortium – St. Louis Missouri

Human Microbiome Project Consortium – September 2010 – St Louis, Missouri

We received warm welcome messages from Dr George Weinstock and Dr Jane Petersen as well as a humorous welcome from Dr Larry Shapiro, Dean of Washington University Medical School. 

It was wonderful to see so many scientists come together to share the progress on their individual HMP related demonstration projects.  Our own demonstration project with Dr Zhiheng Pei, involving the esophagus microbiome and how that relates to esophageal adenocarcinoma (EA), was quite unique compared to the other projects as we were the only group to focus on the correlation between bacterial population and a form of cancer. 

With over 400 participants and 59 speakers, the conference was quite successful and very interesting.  JCVI Director Dr Karen Nelson did a wonderful job moderating one of the segments.  Dr Roger Lasken also gave a thorough presentation on his lab’s single cell approaches to genomic sequencing of uncultureable bacteria.  Johannes Goll gave a great presentation on his recent work with an open source tool called METAREP (recently published in Bioinformatics 8/26/2010), which is designed to help scientists with analyzing annotated metagenomic data.  And Dan Haft presented his interesting work with algorithmically tuning protein families from reference genomes for systems discovery. 

Overall the conference was quite interesting and informative.  I continue to wish all of the participating sequencing centers, PIs, and others involved with the HMP much success with their projects. 

Hope to see everyone in Vancouver!!!

Advance Access JCVI Metagenomics Reports Application Note

A significant JCVI informatics development is JCVI Metagenomics Reports, an open source Web 2.0 application designed to help scientists analyze and compare annotated metagenomics data sets. Users can download the application to upload and analyze their own metagenomics datasets.

METAREP has just been published in Bioinformatics (08/26/2010) as an open access article. The publication is currently accessible under the Bioinformatics Advance Access model. The PDF version can be downloaded at

http://bioinformatics.oxfordjournals.org/cgi/reprint/btq455v1.pdf

Supplementary information includes the METAREP data model and an overview about its search performance accessible at

http://bioinformatics.oxfordjournals.org/cgi/content/full/btq455/DC1

One of METAREP’s  key features that distinguishes it from other metagenomics tools is that it utilizes a high-performance scalable search engine that allows users to analyze and compare extremely large metagenomics datasets, e.g. datasets produced by the Human Microbiome Project.

If you like to learn more about METAREP and talk to the developers,  join us  at  Human Microbiome Research Conference in St. Louis in Missouri (August 31 – September 2, 2010). We will present METAREP  the first day of the conference at 10:35am (see agenda).

Contact Us:

We would like to hear from you. If you have questions or feedback or if you wish to contribute to the METAREP open source project please send an email to metarep-support@jcvi.org

Links:

JCVI’s METAREP Instance

METAREP Flyer

METAREP Manual

METAREP Source Code

High-performance comparative metagenomics

Are your carrying out large scale metagenomics analyses to identify differences among multiple sample sites? Are you looking for suitable analysis  tools?

If you have not yet found the right analysis tool, you may be interested in  the latest beta version of JCVI Metagenomics Reports (METAREP)  [Test It].

METAREP is a new open source tool developed for high-performance comparative metagenomics .

It provides a suite of web based tools to help scientists view, query, browse, and compare metagenomics annotation data derived from ORFs called on metagenomics reads or assemblies.

Users can either specify fields, or logical combinations of fields, to filter
and refine datasets
. Users can compare multiple datasets at various functional and taxonomic levels, applying statistical tests as well as hierarchical clustering, multidimensional scaling, and heatmaps (see image gallery).

For each of these features, tab delimited files can be exported for downstream analysis. The web site is optimized to be user friendly and fast.

Feature Summary [download Flyer]:

  • Handle extremely large datasets. Uses scalable high-performance Solr/Lucene search engine (we have indexed 300 million annotation entries, but much larger volumes can be handled as shown by Hathi Trust).
  • Compare 20+ datasets at the same time. Use various compare
    options including statistical tests and plot options to visualize
    dataset difference at various taxonomic and functional levels.
  • Apply statistical tests such as METASTATS (White et al.), a modified
    non-parametric t-test to compare two sample populations (e.g.
    metagenomics samples from healthy and diseased individuals).
  • Export publication-ready graphics. Export heatmaps, hierarchical clustering, and multi-dimensional scaling plots in PDF format.
  • Analyze KEGG metabolic pathways. Summaries include enzyme
    highlights on KEGG maps, pathway enzyme distributions, and
    statistics about pathway coverage at various pathway levels.
  • Search using a SQL-like query syntax. Build your query using 14
    different fields that can be combined logically.
  • Drill down into data using METAREP’s NCBI Taxonomy, Gene
    Ontology, Enzyme Classification or KEGG Pathway browser.
    Install your own METAREP version.
  • Flexible central configuration, METAREP and 3rd party code base is completely open source.
  • Cross-link function with phylogeny. Slice your data at various
    taxonomic and/or functional levels. For example, search for all
    bacteria or exclude eukaryotes or search for a certain (GO/EC
    ID)/taxonomic combination.
  • Generic data format. Data types that can be populated include a
    free text functional description, best BLAST hit information, as well
    as GO ID, EC ID, and HMMs.

How to analyze your own data: You can install your own METAREP version to analyze your metagenomics annotation data [download source]. We have written a comprehensive manual that describes the installation process step by step [download manual]. Since METAREP only operates on annotated data, raw sequences need to be annotated first. Supported data types that can be loaded for each sequence include functional descriptions, best BLAST hits fields (E-Value, Percent Identity, NCBI Taxon, Percent Sequence Coverage), GO, EC, and HMM assignments. The installation also contains a set of example annotations that can be imported.

Contact Us:

We would like to hear from you. If you have questions or feedback or if you wish to contribute to the METAREP open source project please send an email to metarep-support@jcvi.org

Links:

JCVI’s METAREP Instance

METAREP Flyer

METAREP Manual

METAREP Source Code