Trapping Microbes 750 miles north of the Arctic Circle

About 1% of all microbes are “culturable” in the lab. They are some of the most stubborn organisms requiring special and specific nutrients as well as optimal temperatures and conditions. So, how do we get the “unculturables” to be “culturable”? We make bacteria “traps”, where we take media, sandwich them between a membrane that cannot be penetrated by bacteria and another membrane that can be penetrated by bacteria. Bacteria can grow and migrate toward the trap and essentially get stuck within the membranes and we can cultivate them afterwards. What about bacteria that don’t migrate? We can account for these bacteria by taking them out of the soil, placing them in media and putting them back into the environment using devices called diffusion chambers. These diffusion chambers consist of two membranes that are able to hold bacteria and media yet are porous enough for nutrients and ions. Using these techniques we have already cultivated more than 15 previously uncultivated species. For year two of this project we intend to cultivate at least 90 new species for whole genome shotgun sequencing. WGS data from such new species will be incredibly useful in determining which metabolic pathways are present to allow these organisms to survive in such a harsh barren climate.

Diffusion Chambers

Diffusion Chambers

Setting Bacterial Traps

Setting Bacterial Traps

Traps Under Water

Traps Under Water

Thule, Greenland Year Two

Sequence data from the previous year allowed us to determine the overall microbial population in each site and this year we decided to focus on the Rich Lake site which seem to have representation of nearly all microbes found in the other sites. So lucky for us we only had to work on one site this year rather than six. This in itself had me excited to go back to Thule. After a five-hour flight on a military plane from BWI I finally arrived to Thule Greenland where we were greeted by the Colonel as well as other high ranking military officials at the hanger. Once I cleared the customs processing area, I arrived to the dorm where the other scientists were living. It was a little different from last year’s accommodations but nevertheless the luxuries of WI-FI, Internet and cable TV were all available. As I am anxious to get to the field and see the changes in the Rich Lake site, we were given some interesting news. That day was not a good day to travel to the site because a mother polar bear and her two cubs were spotted nearby not too long ago by military police. However, we managed to get other work done by preparing the schedule for the sampling, cultivation and other labwork.

 

The next few days consisted of preparing culture media, cultivation traps and diffusion chambers, and going out into the field (polar bear spray in hand; yes it’s a real thing!). We were extra careful in the field since there was quite a bit of fog in the area that did not seem to go anywhere and fog happens to be the same color as polar bears. The fog did however make it a bit easier to sleep since most of the sunlight was covered and when there’s 24 hours of daylight from mid-April until September, a little fog can still serve a purpose.

Rich Lake Site

Rich Lake Site

Greenland

Greenland

Scientist Spotlight: Meet Sarah Highlander

Sarah Highlander Ph.D. is an esteemed scientist and professor who joined JCVI in La Jolla this year. She comes from a long line of academically successful Professors, including a great uncle who was a University Dean. As a young child, Sarah was influenced by her parents: her mother was a musician and her father was a Ph.D. chemical engineer. Sarah too was a musician and she still enjoys jazz and the opera. But it was her father’s scientific career that influenced her own decision to pursue scientific research as her career.

Dr. Sarah Highlander

Dr. Sarah Highlander

As a chemical engineer and early IT specialist, he shared his interests with her at the kitchen table by doing mathematical puzzles and simple experiments. They explored the impact light had on grass growth by placing plants in the closet. Then in high school, she had the opportunity to work on a microbiology project with the help of her father. Using agar slants from his colleague’s lab, she looked for antimicrobial features of bacteria in the soil. Even with these opportunities, her focus in the sciences wasn’t fully set until she began working as a technician in a fermentation research lab where she had the opportunity to work with plasmids after completing her bachelor’s degree. At this point, plasmids and restriction enzymes were not readily available and researchers had to isolate them in their labs. She was extremely successful as a technician and even published several papers and secured several patents.

This experience launched Highlander into Medical Microbiology. She went to the Sackler Institute of Biomedical Sciences at the New York University School of Medicine, where she earned her Ph.D. in 1985. With her curious nature and the bourgeoning field of biotechnology, she began to research the replication of DNA plasmids in Staphylococcus. She asked basic but as yet unanswered questions such as, “How are these molecules controlled in the cell?” and “How can they best be manipulated in the laboratory?” Her thesis involved characterizing small RNA molecules that control plasmid copy number.

During her Post-doctoral fellowship, she shifted her focus to infectious diseases and worked on vaccine development for a cattle disease called “shipping fever” at the University of Texas Health Science Center. Shipping fever is the most common and costly problem affecting calves. It accounts for major economic losses to the cattle producer by reducing average daily weight gain, impairs feed efficiency, and diminishes overall performance and health of beef calves. Vaccination is key to reduce the disease and Highlander’s research culminated in the development of a subunit vaccine that is still in use.

After her fellowship, she began her professorship at Baylor’s College of Medicine (BCM), where she continued her research into shipping fever. The primary bacterial agent in this disease is Mannheimia haemolytica, which is the same family as the human respiratory pathogen, Haemophilus influenzae. JCVI scientists were the first to sequence and publish the H flu genome in 1995. Dr. Highlander’s group performed extensive characterization of the M. haemolytica leukotoxin and developed numerous genetic tools for manipulation and tagging of the organism. She holds patents for subunit and live-attenuated vaccines to prevent shipping fever.

In 2002, Highlander founded Prokaryon Technologies, a for-profit company focused on animal health to prevent and control diseases associated with food animals. One of Prokaryon’s lead products was a genomics-derived vaccine to prevent shipping fever in cattle.

While leading and growing her company, Highlander stayed committed to her academic research interests and joined the Human Genome Sequencing Center at Baylor. At BCM, she participated in genome sequencing of several pathogens (including M. haemolytica) and she moved to focus more on human pathogens. From 2006 to 2013, Highlander was a principal investigator for the Human Microbiome Project (HMP), a National Institutes of Health-funded program in which JCVI researchers were also key leaders.

In addition to her research, Highlander was involved in graduate and medical education at BCM. She was the co-director of the departmental graduate program for 15 years and directed and taught courses focused on bacterial physiology and molecular laboratory methods. Preparation for lectures and interactions with students helped her stay on top of new techniques and research, which in turn helped her further her own research. Sarah had the opportunity to mentor many graduate students both formally and informally.

At JCVI, Highlander is continuing her work on the microbes that live in and on the human body. Specifically she and her team are looking at the complex microbial communities that live in the human gut. While many microbes are associated with disease, most in the human body are associated with health. Highlander and her team are working to develop specific healthy bacterial mixtures that could be used treat conditions such recurrent Clostridium difficile diarrhea, inflammatory bowel disease and others. She is also using bioinformatics tools to look for new causes of diarrhea. “I am delighted to be a part of the collaborative environment here at JCVI and to be surrounded by colleagues who share common interests in bacterial genetics, genomics, microbial physiology and pathogenesis. The microbiome group at JCVI is strong and I hope to be able to make significant contributions to ongoing and future projects here”.

Even in her personal life, Sarah researches, through her hobby of tracing her genealogy. She has been able to find family roots dating back to the 1500s. This detective work is challenging but it keeps her mind sharp and detailed oriented. She points out that learning family naming patterns can be critical to genealogy research just as algorithm development is to genomic research.

Never having lost that early scientific curiosity and excitement of discovery that her father instilled in her as a young girl, Sarah loves working in the laboratory at JCVI and asking questions. Her analytical and inquisitive nature is one of her greatest professional strengths. She is fascinated by the complexity of the microbial ecosystem in our bodies and the impact these microbes have on our health. As she says, “Microbes are going to continue to win through evolution. We need to figure out the next step to keep ahead!” Let’s hope Highlander and her team can win this battle.

Professional Development Opportunities this Summer

This summer we are offering two professional development workshops: GenomeSolver and Bioinformatics: Unlocking Life through Computation.  Both explore bioinformatics, microbial diversity and the implementation in the undergradauate or high school classrooms. 

The GenomeSolver workshop trains faculty on genome analysis. Workshop attendees will learn about general methodologies, standards, and processes used to annotate and analyze microbial genomes. The workshop contents will be available to aid the faculty in developing teaching modules. In addition, extensive documentation on methodologies and tools will be available via the online environment created for this project. On online web portal Genome Solver (www.genomesolver.org) will be a virtual space for development and sustaining of community. Genome Solver will assist faculty with technical issues and curricular design, as well as an online environment for the ongoing sharing of information including publication of student work. 

http://www.jcvi.org/cms/education/prodev/genome-solver-annotation-workshops

Bioinformatics: Unlocking Life through Computation is a new opportunity for high school teachers. Genomics and biotechnology are valuable tools in our quest to understand life and nature. However, introducing the science classroom to the computational and mathematical underpinnings of biology can be challenging. The goal of this workshop is to introduce a curriculum for mathematics and science education in the area of genomics (with a focus on the fascinating world of microbes). Educators will be introduced to the various analysis and computational challenges that arise in this discipline. Workflow examples illustrating comparative genomic analysis will be made available through the JCVI Metagenomics Report (METAREP) software infrastructure. The eventual aim is for the educational material to be integrated with local high school curricula requirements to expose students to both hypothesis-driven and discovery-based science.

 http://www.jcvi.org/cms/education/prodev/bioinformatics-unlocking-life-through-computation/

JCVI Hosts South African Scientists to Share Microbiome Research Techniques

Two scientists from the University of Cape Town, South Africa have joined Dr. Bill Nierman’s lab for the next month as part of NIH’s Human Heredity and Health in Africa (H3Africa) Initiative, a training program designed to build out technical biological skills in the African research community. This training relates specifically to developing techniques around the area of microbiome analysis, a relatively new field in the biological sciences.

Microbiome analysis for the collaborative study is looking at entire community of microorganisms in the respiratory tract of South African infants to better understand how the microbiome is associated with infant pneumonia and wheezing episodes. The expectation is that the organisms that reside in the infant respiratory tract will provide protection from or a predisposition to the pneumonia or wheezing episodes.

 

The Nierman Group

The Nierman group left to right Sarah Lucas, Bill Nierman, Shantelle Claassen, Mamadou Kaba and Stephanie Mounaud (unpictured Jyoti Shanker and Lilliana Losada) welcomes visiting scientists Ms. Classeen and Dr. Kaba from University of Cape Town for a month long training in microbiome sequencing and analysis.

Mamado Kaba, MD, PhD and colleague Shantelle Claassen from the University of Cape Town will be working closely under the guidance of JCVI’s Stephanie Mounaud who is functioning as the project manager and coordinating the laboratory components of a similar project at JCVI studying the microbiomes of inafnts in the Philippines and also in South Africa. These studies are sponsored by the Bill and Melinda Gates Foundation. The training will focus initially on preparing samples for DNA sequencing on a modern DNA sequencing platform, the Illumina MiSeq instrument. Once the sequence reads are off the sequencer, the instructional focus will shift to analysis of the reads by means of an informatics pipeline that develop phylogenies, or family trees, of the microbes that are obtained from the infant respiratory tract so that the abundance and relatedness of the microbes can be established. The bioinformatics training will be provided by Jyoti Shankar, the statistical analyst working on the Gates Foundation Project.

Mamadou Kaba is a Wellcome Trust Fellow working in the Division of Medical Microbiology, Faculty of Health Sciences, University of Cape Town. Mamadou’s research interests include the molecular epidemiology of infectious diseases and the study of human microbiome in healthy and disease conditions. He has contributed in establishing a new research group conducting studies on how the composition of the upper respiratory tract, gastrointestinal, and the house dust microbial communities influences the development of respiratory diseases.

Prior to joining the University of Cape Town, Mamadou worked as Research Associate at the Laboratory of Medical Microbiology, Timone University Hospital, Marseille, France, where he studied the epidemiological characteristics of infection with hepatitis E virus in South-eastern France.

Shantelle Claassen is pursuing a Masters degree in the Division of Medical Microbiology at the University of Cape Town. She has completed a BSc (Med) Honours degree in Infectious Diseases and Immunology at the University of Cape Town, during which she examined the relative efficacy of extracting bacterial genomic DNA from human faecal samples using five commercial DNA extraction kits. The DNA extraction kits were evaluated based on their ability to efficiently lyse bacterial cells, cause minimal DNA shearing, produce reproducible results and ensure broad-range representation of bacterial diversity.

Mamadou and Shantelle are currently involved in an additional prospective, longitudinal study of which the primary objective is to investigate the association between fecal bacterial communities and recurrent wheezing during the first two years of life.

Building the World’s First Net-Zero Energy Lab [video]

Building the World’s First Net-Zero Energy Lab

And see the construction in time-lapes.

Amazon Expedition

Yesterday, JCVI expedition scientist Jeff Hoffman embarked from Manaus on a sampling expedition of the Amazon River and its tributaries, which contains 1/5th of the Earth’s river flow. In collaboration with scientists Dr. Guilherme Oliviera and Dr. Sara Cuadros from the Centro de Excelencia em Bioinformatica (CEBIO) of Belo Horizonte, Jeff is taking samples to characterize the genomes of microbes found along 2/3rds of the entire Amazon watershed, including inflowing rivers from Manaus to Macapa. Our collaborators at CEBIO will be sequencing the samples with a joint Brazil-USA effort on analysis. Long recognized for the biodiversity of visible organisms, the Amazon is understudied with regards to the diversity of microscopic organisms and this expedition will substantially increase our understanding of the biological diversity on Earth. This work continues, leverages, and complements previous and ongoing JCVI work characterizing the unexplored microbiomes of marine, estuarine, freshwater, and terrestrial environments around the world.

See a gallery of the expedition on Facebook. More pictures will be added throughout the trip.

The 2014 Summer Internship Application is Open and Announcing the Genomics Scholar Program

The 2014 Summer Internship Application is now open.   Last summer, we hosted 49 interns from a pool of 424 applicants. They presented their research in the First Annual Summer Internship Poster Sessions held in San Diego and Rockville. The posters were judged by a team of volunteer JCVI scientists and the poster sessions were open to all employees, interns and their guests to share what great work they all participated in this summer.

 

 

2013 Intern Poster Session

2013 Intern Poster Session

We are also excited to announce the new Genomics Scholar Program beginning this summer and also accepting applications.  The Genomic Scholar Program (GSP) is a targeted research experience program to community college students in Rockville. Our program incorporates multiple avenues of support for students through the research experience with the Principal Investigators as mentors, and supplemental professional development provided by the JCVI.  Additionally, selected students will have the opportunity to participate in undergraduate research conferences.

The GSP is supported by the National Institute of Diabetes and Digestive and Kidney Diseases of the National Institutes of Health under award number R25DK098111.

Sampling: US to the Azores

I’m off again on an ocean sampling voyage but this time instead of being onboard the JCVI’s Sorcerer II, I am onboard the R/V Endeavor as part of a multi-institution, international scientific sampling team that is headed from the US to the Azores.

On Thursday August 22 we left Morehead City, North Carolina for Ponta Delgada on Sao Miguel Island in the Azores.  The research vessel will take multiple samples along the 23 day transect.  Here is a rundown of the teams and the science we are conducting.

Crew leaving Morehead City, NC.

Crew leaving Morehead City, NC. From the left: Sarah Fawcett, Amandine Sabadel, Malcolm Woodward, and Bess Ward.

R/V Endeavor

R/V Endeavor

I will be filtering large volumes of seawater on 293mm filters for DNA sequencing, as well as smaller volumes onto smaller Sterivex filters for RNA sequencing and associated studies of gene expression within various microbial communities. This research expedition is funded by a grant from the National Science Foundation program in Dimensions of Biodiversity to Bess Ward at Princeton University and Andrew Allen at JCVI. The goal of our JCVI group is to extend findings from the Sorcerer II Global Ocean Sampling program, which documented massive genomic diversity and unusual physiological and biochemical capabilities within and between many lineages of marine microorganism. With samples collected on this research cruise, we will have the opportunity to document large-scale patterns in gene expression, and generate key hypotheses related to the most biochemically-active microbes across a major section of the upper 1000m of the North Atlantic. Data obtained from this study will be combined with similar data we collected last February and August on cruises out of Bermuda to the Bermuda Atlantic Time Series (BATS) stations in the in the sub-tropical Atlantic.

North Atlantic Transect, north of Sorcerer II transect to the Azores in 2009.

North Atlantic Transect, north of Sorcerer II transect to the Azores in 2009.

The Princeton team headed up by Bess Ward includes Sarah Fawcett, Nicolas Van Oostende, Jess Lueders-Dumont, Dario Marconi, and Keiran Swart. Their primary research involves using flow cytometry to physically capture, size fractionate and identify microbes living in the sunlit layer of the ocean. These microbes are directly responsible for assimilation of dissolved nitrate, which accumulates in the dark interior of the ocean. Specific identification of these microbes is an important research goal for microbial oceanography because the regulation and magnitude of global oceanic CO2 assimilation is driven explicitly by nitrate assimilation by photosynthetic microbes. Such microorganisms also produce a large fraction of the oxygen in the atmosphere. The Princeton group will perform nitrification experiments and measure levels of dissolved nitrate, ammonia and carbon by using stable and natural isotope tracers. The team will investigate the origins of dissolved inorganic nitrogen by measuring the natural abundance of the nitrogen isotopes.  Net tows will also be performed to collect the “bigger” planktonic organisms, such as zooplankton, within the ocean food chain.

Real time nutrient data down to nanomolar levels will be determined by Malcolm Woodward of Plymouth Marine Laboratory (PML) and Amandine Sabadel from the University of Otago in New Zealand.

As we motor to our first station, which we should reach on Monday September 2nd, we stop every morning at 5 am to perform a CTD cast to 1000 meters.  Based on biological and physical features, observable in real time via CTD sensors cabled to the shipboard computer,12 bottles, each containing 30 liters of sea water, are sealed at varied depths and the 360 liters is brought to the boats deck.  Once the CTD is on the deck, the different scientists scurry to gather their allocated amount of water from the CTD rosette and hurry back to their labs to do the appropriate work.

CTD Controls

CTD Controls

CTD Controls

CTD Controls

CTD1

CTD1

As of Wednesday August 28, 2013, we have done 7 transect CTD casts, all but one to 1000 meters.  Today we sampled on the Grand Banks and the water column depth was only 57 meters. For every cast I have collected RNA samples at 1000 meters, 250 meters, within the Deep Chlorophyll Max (DCM) (if no DCM is apparent, then just below the Chlorophyll max), a sample from within the Chlorophyll max and in the mixed layer (normally at 20 meters).

The weather has been great except for one 24 hour period when the swells grew to about 7 feet and the boat was really rolling back and forth.  The crew is great, the food is awesome, good thing they have a small gym or I don’t think most of us would fit in our clothes after a few weeks out here! The scientists are working well as a team and this should be a very exciting and beneficial science expedition.

CTD Profile

CTD Profile

Dry Lab

Dry Lab

 

Once we get to the our first station we will stay there for two days………….it will be a very intense two days, then a day motor to the second station followed by another crazy two days of sampling………….more on that next blog!

Thule, Greenland – Day One

Arrived at Thule, Greenland after a 5 hr flight from Copenhagen.  It was pretty interesting seeing a long line of people all getting on a flight that was headed to a part of the world that usually has less than 600 people there at any given time.  Arrival was pretty straightforward, no jetway, no customs, no LCD screens telling you where to pick up your bag.  Just a few military personnel checking your documents to ensure that you have the approval from the Danish government and USAF to be on base.  First impression getting off the plane…it’s cold.  Not as cold as I expected it to be but it was just 90 degrees F when I left home a few days ago.  Today’s high was 39 degrees F.  Standing in the sun it’s not so bad but when the wind starts blowing it turns into a recipe for chapped lips and windburn.  Oh and did I mention the massive mosquitos here?  Not much wildlife in this part of the world but the mosquitos outnumbers the vertebrates probably a million to one.  They are also VERY aggressive; they even swarmed the trucks while we were driving around the base.  We were shown our living quarters, which were very nice, kind of reminded me of living in the dorms during undergrad.  There are individual rooms and a shared bathroom on each floor.  We toured the various sites that our collaborator Slava Epstein already pointed out as good sampling sites that vary in vegetation and proximity to water.  The land here is quite desolate, not much green, mostly moss and small shrubs growing.  Traditional trees are nonexistent but “ground trees” are actually common.  They are trees that grow outward on the grass and not upward.  The rest resembles pictures taken by the mars rover.  As the day goes by I noticed the sun was circling and I came to the realization that the typical artic summer was happening right in front of me.  The sun literally circles and will not go down until around September.  It was quite odd, getting in bed at midnight and seeing the sun still in the sky.  Tomorrow will be more interesting since we will be going further away from base to sample additional areas. 

blog2

blog1