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 One step closer to reality
20.04.2018 | Max-Planck-Institut für Entwicklungsbiologie

nachricht The dark side of cichlid fish: from cannibal to caregiver
20.04.2018 | Veterinärmedizinische Universität Wien

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

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

Im Focus: Spider silk key to new bone-fixing composite

University of Connecticut researchers have created a biodegradable composite made of silk fibers that can be used to repair broken load-bearing bones without the complications sometimes presented by other materials.

Repairing major load-bearing bones such as those in the leg can be a long and uncomfortable process.

Im Focus: Writing and deleting magnets with lasers

Study published in the journal ACS Applied Materials & Interfaces is the outcome of an international effort that included teams from Dresden and Berlin in Germany, and the US.

Scientists at the Helmholtz-Zentrum Dresden-Rossendorf (HZDR) together with colleagues from the Helmholtz-Zentrum Berlin (HZB) and the University of Virginia...

Im Focus: Gamma-ray flashes from plasma filaments

Novel highly efficient and brilliant gamma-ray source: Based on model calculations, physicists of the Max PIanck Institute for Nuclear Physics in Heidelberg propose a novel method for an efficient high-brilliance gamma-ray source. A giant collimated gamma-ray pulse is generated from the interaction of a dense ultra-relativistic electron beam with a thin solid conductor. Energetic gamma-rays are copiously produced as the electron beam splits into filaments while propagating across the conductor. The resulting gamma-ray energy and flux enable novel experiments in nuclear and fundamental physics.

The typical wavelength of light interacting with an object of the microcosm scales with the size of this object. For atoms, this ranges from visible light to...

Im Focus: Basel researchers succeed in cultivating cartilage from stem cells

Stable joint cartilage can be produced from adult stem cells originating from bone marrow. This is made possible by inducing specific molecular processes occurring during embryonic cartilage formation, as researchers from the University and University Hospital of Basel report in the scientific journal PNAS.

Certain mesenchymal stem/stromal cells from the bone marrow of adults are considered extremely promising for skeletal tissue regeneration. These adult stem...

Im Focus: Like a wedge in a hinge

Researchers lay groundwork to tailor drugs for new targets in cancer therapy

In the fight against cancer, scientists are developing new drugs to hit tumor cells at so far unused weak points. Such a “sore spot” is the protein complex...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

VideoLinks
Industry & Economy
Event News

Invitation to the upcoming "Current Topics in Bioinformatics: Big Data in Genomics and Medicine"

13.04.2018 | Event News

Unique scope of UV LED technologies and applications presented in Berlin: ICULTA-2018

12.04.2018 | Event News

IWOLIA: A conference bringing together German Industrie 4.0 and French Industrie du Futur

09.04.2018 | Event News

 
Latest News

Magnetic nano-imaging on a table top

20.04.2018 | Physics and Astronomy

Start of work for the world's largest electric truck

20.04.2018 | Interdisciplinary Research

Atoms may hum a tune from grand cosmic symphony

20.04.2018 | Physics and Astronomy

VideoLinks
Science & Research
Overview of more VideoLinks >>>