Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

TraDIS technique tackles typhoid

20.10.2009
First high-throughput analysis of every Salmonella Typhi gene

For the first time, researchers are able to look at the need for every gene in a bacterial cell in a single experiment. The new method will transform the study of gene activity and the search for weaknesses in bacterial armouries.

Using a newly developed, next-gen sequencing method, a team established which genes Salmonella Typhi needs to survive and which are more of a luxury. The results and the method will be a boon to scientists tackling bacterial disease, allowing them to capitalize on the abundance of genomic sequence data from next-generation sequencing technologies.

Every year 22 million people are infected and 220,000 die from infection with S. Typhi. It is a special threat in the developing world, in areas with poor sanitation or a lack of clean drinking water.

The team were able to look at almost all the genes in S. Typhi and showed that it needs only 356 genes for survival: 4162 genes were not essential. Knowing which genes are essential to the survival of pathogens, researchers can seek treatments to target those genes.

"We developed a new method that is ten times more powerful than any previous technique," says Sanger Institute graduate student Gemma Langridge, one of the first authors on the paper. "By combining transposon-induced mutagenesis – a method whereby small chunks of cut-and-paste DNA sequence are inserted into the genome effectively disabling individual genes – and high-throughput sequencing, we have been able to determine which genes are essential for the survival of S. Typhi and which are non-essential."

"Crucially, our new method allows us to achieve all this in just a single experiment."

Using the novel method, which the team have named TraDIS (Transposon Directed Insertion site Sequencing), they inserted transposons into the S. Typhi genome to generate more than one million mutants. They then grew the bacteria and used next-generation sequencing to directly identify 370,000 insertion sites in the S. Typhi genome – an average of more than 80 insertion sites per gene. Previous methods produce only a few mutations per gene.

If a transposon inserts into an essential gene, the gene is silenced and that mutant cell will not grow and it – and the transposon insert – will be absent from the mutant pool. By sequencing DNA from the entire pool – approximately 1 million mutants in total – the team were able to identify genes in which no transposon insertions had been detected.

In a single experiment using the TraDIS method, the team were able to determine whether or not 99.6% of the S. Typhi genes are essential to its survival.

"Sequencing centres such as ours can produce vast amounts of genomic data at a pace unimaginable just a few years ago," explains Professor Julian Parkhill, Director of Sequencing and head of Pathogen Genomics at the Sanger Institute. "One of our aims is to develop high-throughput research methods that can exploit this explosion of genetic data, to ensure these resources can be used effectively. We can now discover which of all the genes in an organism are essential to its survival or required for growth under special conditions, such as infection. Our new TraDIS method will make a dramatic difference to the ability to carry out such genome-wide research."

Importantly, the team applied the method to a clinical problem by looking at how S. Typhi might survive in humans. Typhoid can be spread by carriers who, without showing symptoms, act as reservoirs, storing the bacterium in the gallbladder and passing it to others. The most famous such carrier was Typhoid Mary, who worked in the food industry in the US and spread typhoid fever without exhibiting any symptoms herself.

But, bacteria cannot survive in the fairly hostile environment of the gall bladder unless they are tolerant to bile – the fatty fluid secreted by the gall bladder. Looking at genes involved in bile resistance, allows us to see which genes are essential for helping S. Typhi persist in a carrier.

"We grew the bacteria in ox bile to pick out genes required for bile tolerance," says Keith Turner, Sanger Institute investigator and a senior author on the paper. "We found 169 genes involved in bile tolerance – many of these had not been suspected before and more than 30 are genes not characterized at all.

"Using TraDIS, we have highlighted several possible new targets for treatment that would pick on S. Typhi's need to survive in the gall bladder."

For the first time, it is possible to paint a comprehensive picture of essential, advantageous or burdensome genes in many phases of the bacterial life cycle, to determine functions necessary to support them throughout their entire disease cycle. Such a picture is important for discovery of new targets for treatment.

This elegant new method exemplifies how high-throughput research allows scientists to determine systematically the function of or requirement for individual genes in a single experiment, opening the door for similar analyses of other pathogenic genomes in the future.

Publication Details
Langridge G C, Phan M-D, Turner D J et al. (2009) Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants. Genome Research

Published online before print at doi: 10.1101/gr.097097.109

Funding

This work was supported by the Wellcome Trust.

Participating Centres

The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, UK
Environmental Research Institute, University College, Lee Road, Cork, Ireland
Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield, UK
Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, UK

Laboratory of Gastrointestinal Pathogens, Centre for Infections, Health Protection Agency, Colindale, London, UK

The Wellcome Trust Sanger Institute, which receives the majority of its funding from the Wellcome Trust, was founded in 1992. The Institute is responsible for the completion of the sequence of approximately one-third of the human genome as well as genomes of model organisms and more than 90 pathogen genomes. In October 2006, new funding was awarded by the Wellcome Trust to exploit the wealth of genome data now available to answer important questions about health and disease. http://www.sanger.ac.uk

The Wellcome Trust is the largest charity in the UK. It funds innovative biomedical research, in the UK and internationally, spending over £600 million each year to support the brightest scientists with the best ideas. The Wellcome Trust supports public debate about biomedical research and its impact on health and wellbeing. http://www.wellcome.ac.uk

Don Powell | EurekAlert!
Further information:
http://www.sanger.ac.uk

More articles from Life Sciences:

nachricht Researchers identify potentially druggable mutant p53 proteins that promote cancer growth
09.12.2016 | Cold Spring Harbor Laboratory

nachricht Plant-based substance boosts eyelash growth
09.12.2016 | Fraunhofer-Institut für Angewandte Polymerforschung IAP

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Electron highway inside crystal

Physicists of the University of Würzburg have made an astonishing discovery in a specific type of topological insulators. The effect is due to the structure of the materials used. The researchers have now published their work in the journal Science.

Topological insulators are currently the hot topic in physics according to the newspaper Neue Zürcher Zeitung. Only a few weeks ago, their importance was...

Im Focus: Significantly more productivity in USP lasers

In recent years, lasers with ultrashort pulses (USP) down to the femtosecond range have become established on an industrial scale. They could advance some applications with the much-lauded “cold ablation” – if that meant they would then achieve more throughput. A new generation of process engineering that will address this issue in particular will be discussed at the “4th UKP Workshop – Ultrafast Laser Technology” in April 2017.

Even back in the 1990s, scientists were comparing materials processing with nanosecond, picosecond and femtosesecond pulses. The result was surprising:...

Im Focus: Shape matters when light meets atom

Mapping the interaction of a single atom with a single photon may inform design of quantum devices

Have you ever wondered how you see the world? Vision is about photons of light, which are packets of energy, interacting with the atoms or molecules in what...

Im Focus: Novel silicon etching technique crafts 3-D gradient refractive index micro-optics

A multi-institutional research collaboration has created a novel approach for fabricating three-dimensional micro-optics through the shape-defined formation of porous silicon (PSi), with broad impacts in integrated optoelectronics, imaging, and photovoltaics.

Working with colleagues at Stanford and The Dow Chemical Company, researchers at the University of Illinois at Urbana-Champaign fabricated 3-D birefringent...

Im Focus: Quantum Particles Form Droplets

In experiments with magnetic atoms conducted at extremely low temperatures, scientists have demonstrated a unique phase of matter: The atoms form a new type of quantum liquid or quantum droplet state. These so called quantum droplets may preserve their form in absence of external confinement because of quantum effects. The joint team of experimental physicists from Innsbruck and theoretical physicists from Hannover report on their findings in the journal Physical Review X.

“Our Quantum droplets are in the gas phase but they still drop like a rock,” explains experimental physicist Francesca Ferlaino when talking about the...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

ICTM Conference 2017: Production technology for turbomachine manufacturing of the future

16.11.2016 | Event News

Innovation Day Laser Technology – Laser Additive Manufacturing

01.11.2016 | Event News

#IC2S2: When Social Science meets Computer Science - GESIS will host the IC2S2 conference 2017

14.10.2016 | Event News

 
Latest News

Researchers identify potentially druggable mutant p53 proteins that promote cancer growth

09.12.2016 | Life Sciences

Scientists produce a new roadmap for guiding development & conservation in the Amazon

09.12.2016 | Ecology, The Environment and Conservation

Satellites, airport visibility readings shed light on troops' exposure to air pollution

09.12.2016 | Health and Medicine

VideoLinks
B2B-VideoLinks
More VideoLinks >>>