Posts Tagged ‘DNA’

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UPC teams conduct research in biomedical engineering to improve people’s health

UPC teams conduct research in biomedical engineering to improve people’s health

Systems to improve patient rehabilitation, methods that help detect diseases, and smart biomaterials for optimising treatments—scientific advances in the field of biomedical engineering are unstoppable. A number of leading UPC teams are carrying out research aimed at harnessing technology to improve people’s health.

Parkinson’s disease is the second most common neurodegenerative disease after Alzheimer’s. Optimising treatment and rehabilitation of the people it affects and improving their quality of life is the goal of Joan Cabestany and Andreu Català, researchers at the Technical Research Centre for Dependency Care and Autonomous Living (CETpD) of the Universitat Politècnica de Catalunya · BarcelonaTech (UPC).

The two engineers are heading up a European project known as REMPARK (Personal Health Device for the Remote and Autonomous Management of Parkinson’s Disease), which has a budget of €4.73 million. The objective is to develop a pioneering wearable monitoring system that can be used to identify and quantify, in real time and with high reliability, the motor status of Parkinson’s patients during their everyday lives. The system will act automatically—though always under medical supervision—in response to the situations that are most incapacitating for patients, intervening in the least invasive and most effective way possible. Other participants in this ambitious project coordinated by the UPC include the Teknon Medical Centre, Telefónica R&D, the European Parkinson’s Disease Association, and a number of research centres and companies based in Germany, Portugal, Italy, Israel, Ireland, Sweden and Belgium.

The system being developed consists of two elements: a bracelet equipped with a sensor for measuring tremor in patients and a smart device the size of a mobile phone, which is worn at the waist on a belt made of biocompatible material. The device is equipped with a set of sensors and has the capacity to process and wirelessly transmit all the information collected and processed.

When a gait-freezing episode occurs, the REMPARK system will act to synchronise the patient’s movements. This will be achieved by means of auditory, visual or haptic (touch-related) cueing devices, a pump for regulated subcutaneous drug delivery, and a functional electrical stimulation (FES) system. “The device will make it possible to quantify the effects of a drug in a particular patient and adjust the dose accordingly,” says Joan Cabestany, stressing that REMPARK is “a personalised system that adapts to each person’s needs.”

For the first time in Europe, REMPARK will be tested on a hundred patients in their homes. “We want to use the technology to give Parkinson’s patients back their confidence, which is gradually eroded by the disease,” says Andreu Català. The project “will reduce the number of hospitalisations and improve patient treatment and rehabilitation,” adds the researcher, who works at the Vilanova i la Geltrú Campus.

Stress-free cells

The REMPARK project is set to run until 2015, but others are yielding results that are about to hit the market. This was clear at the BIO International Convention, the world’s largest biotechnology exhibition, which was held in Boston (Massachusetts, United States) last June.

The UPC presented a number of patents at the event, including an automatic method for introducing substances such as drugs and DNA into cells (transfection). The method, known as in vitro electroporation, is more efficient and economical than existing approaches.

The technique, which is applied manually, is commonly used in gene therapy, cell-based therapies, and tumour treatment by electrochemotherapy. Cells are detached from the bottom of the plates where they are grown and put into suspension, i.e. into a mixture. They are then placed in a special cuvette with aluminium electrodes on its sides. The cuvette is loaded into a device (an electroporator) that creates a high-intensity electric field across the cells, causing the pores in the cell membrane to open. Substances can then be introduced through these pores.

The new system simplifies and automates this process. A microelectrode assembly is introduced directly into the culture plate and placed at a distance of 10 ìm (10 millionths of a metre) from the cells. A 20 V electric field is then applied (in the conventional process a 500 V field is used). The lower voltage reduces the cost of the devices used to carry out these biotechnological processes and subjects the cells to less stress. The low cost of the microelectrodes also makes it possible to produce single-use electroporators. This patent was developed by researcher Ramon Bragós and doctoral student Tomàs Garcia, who are attached to the Biomedical Engineering Research Centre (CREB), in collaboration with a team at the University of Barcelona (UB).

The UPC is also contributing to major advances in the development of medical devices and diagnostic imaging. The UPC’s Institute of Industrial and Control Engineering (IOC) and the Pulmonology Research Group of Bellvitge Hospital’s Institute for Biomedical Research have developed a virtual bronchoscopy system that improves the diagnosis of lung cancer. The technology provides doctors with information that enables them to decide with more confidence whether an actual bronchoscopy is necessary or not. This helps minimise risk and discomfort for patients.

The system is based on images provided by a virtual bronchoscopy using 2D computed tomography images. The novel feature of the system is that it takes into account the geometry and kinematic constraints of the bronchoscope.

The device is designed so that a pulmonologist can virtually navigate through a patient’s airways and simulate the movements that will later be executed when a flexible bronchoscope is used to perform the examination. It is a useful tool that facilitates “very realistic planning of the most feasible path from the trachea to peripheral pulmonary lesions,” says Jan Rosell, the researcher who carried out the project together with Paolo Cabras and Alexander Pérez, who also work with the IOC. “Doctors can also use the device to determine whether the end of the bronchoscope will reach a lesion, or, if not, how close it can be manoeuvred and what technique will need to be used to obtain a biopsy sample,” Rosell adds.

In addition to pursuing advances in diagnostic imaging, molecular biology and telemedicine, UPC researchers are also doing innovative work in another area of interest: metabolomics, the scientific study of chemical processes involving metabolites. It is in this field that another CREB team has patented an innovative software tool. The advanced program, based on a new algorithm, helps medical professionals make more accurate, automated predictions in disease diagnosis and drug screening.

Developed by Àlex Perera and Francesc Fernández in collaboration with researchers with the University of Barcelona’s Department of Nutrition and Food Science, the tool improves detection of biomarkers, the biological markers used to detect diseases.

Another advantage of the software is that it reduces prediction error in metabolomic analysis and testing (used to examine the small organic molecules in biological systems). Metabolomic analyses are based on biological samples of urine or blood, nuclear magnetic resonance (NMR) techniques, and mass spectrometry (LC/MS). Making predictions in this area is complex because it requires analysis of extensive data obtained from individual samples, but it is of vital importance in evaluating the effectiveness of new drugs, for example.

New test for tuberculosis

Tuberculosis is one of the diseases that accounts for the most morbidity and mortality worldwide. Despite this, there are still a lot of unanswered questions about the disease and many scientific challenges remain to be tackled. Daniel López Codina and Clara Prats of the UPC’s Discrete Modelling and Simulation of Biological Systems group have carried out research in this field in collaboration with a team at the Experimental Tuberculosis Unit of the Germans Trias i Pujol Health Sciences Research Institute Foundation.

The two teams have patented a new method that offers a fast, easy and reliable way to determine the virulence (ability to produce disease) of Koch’s bacillus. The technique allows specialists to make more accurate diagnoses.

López Codina’s team observed the tuberculosis bacillus (Mycobacterium tuberculosis) in an in vitro culture and looked at the way it grows by forming clumps. Given the difficulty of applying conventional microbiological methods with this type of culture, the researchers used an alternative approach: microscopy and analysis with image processing techniques. “This is the first time we’ve been able to use a culture to observe two different strains of the bacterial parasite and the existence of a correlation between the characteristic clumping pattern and the virulence of the disease,” said the researcher.

The results have created a new business opportunity for companies involved in biomedical imaging and diagnostic testing.

Projects like these highlight the huge potential of engineering and medicine to continue delivering solutions that improve people’s quality of life.

Source : http://www.news-medical.net/news/20121123/UPC-teams-conduct-research-in-biomedical-engineering-to-improve-peoples-health.aspx

Full story

UPC teams conduct research in biomedical engineering to improve people’s health

UPC teams conduct research in biomedical engineering to improve people’s health

Systems to improve patient rehabilitation, methods that help detect diseases, and smart biomaterials for optimising treatments—scientific advances in the field of biomedical engineering are unstoppable. A number of leading UPC teams are carrying out research aimed at harnessing technology to improve people’s health.

Parkinson’s disease is the second most common neurodegenerative disease after Alzheimer’s. Optimising treatment and rehabilitation of the people it affects and improving their quality of life is the goal of Joan Cabestany and Andreu Català, researchers at the Technical Research Centre for Dependency Care and Autonomous Living (CETpD) of the Universitat Politècnica de Catalunya · BarcelonaTech (UPC).

The two engineers are heading up a European project known as REMPARK (Personal Health Device for the Remote and Autonomous Management of Parkinson’s Disease), which has a budget of €4.73 million. The objective is to develop a pioneering wearable monitoring system that can be used to identify and quantify, in real time and with high reliability, the motor status of Parkinson’s patients during their everyday lives. The system will act automatically—though always under medical supervision—in response to the situations that are most incapacitating for patients, intervening in the least invasive and most effective way possible. Other participants in this ambitious project coordinated by the UPC include the Teknon Medical Centre, Telefónica R&D, the European Parkinson’s Disease Association, and a number of research centres and companies based in Germany, Portugal, Italy, Israel, Ireland, Sweden and Belgium.

The system being developed consists of two elements: a bracelet equipped with a sensor for measuring tremor in patients and a smart device the size of a mobile phone, which is worn at the waist on a belt made of biocompatible material. The device is equipped with a set of sensors and has the capacity to process and wirelessly transmit all the information collected and processed.

When a gait-freezing episode occurs, the REMPARK system will act to synchronise the patient’s movements. This will be achieved by means of auditory, visual or haptic (touch-related) cueing devices, a pump for regulated subcutaneous drug delivery, and a functional electrical stimulation (FES) system. “The device will make it possible to quantify the effects of a drug in a particular patient and adjust the dose accordingly,” says Joan Cabestany, stressing that REMPARK is “a personalised system that adapts to each person’s needs.”

For the first time in Europe, REMPARK will be tested on a hundred patients in their homes. “We want to use the technology to give Parkinson’s patients back their confidence, which is gradually eroded by the disease,” says Andreu Català. The project “will reduce the number of hospitalisations and improve patient treatment and rehabilitation,” adds the researcher, who works at the Vilanova i la Geltrú Campus.

Stress-free cells

The REMPARK project is set to run until 2015, but others are yielding results that are about to hit the market. This was clear at the BIO International Convention, the world’s largest biotechnology exhibition, which was held in Boston (Massachusetts, United States) last June.

The UPC presented a number of patents at the event, including an automatic method for introducing substances such as drugs and DNA into cells (transfection). The method, known as in vitro electroporation, is more efficient and economical than existing approaches.

The technique, which is applied manually, is commonly used in gene therapy, cell-based therapies, and tumour treatment by electrochemotherapy. Cells are detached from the bottom of the plates where they are grown and put into suspension, i.e. into a mixture. They are then placed in a special cuvette with aluminium electrodes on its sides. The cuvette is loaded into a device (an electroporator) that creates a high-intensity electric field across the cells, causing the pores in the cell membrane to open. Substances can then be introduced through these pores.

The new system simplifies and automates this process. A microelectrode assembly is introduced directly into the culture plate and placed at a distance of 10 ìm (10 millionths of a metre) from the cells. A 20 V electric field is then applied (in the conventional process a 500 V field is used). The lower voltage reduces the cost of the devices used to carry out these biotechnological processes and subjects the cells to less stress. The low cost of the microelectrodes also makes it possible to produce single-use electroporators. This patent was developed by researcher Ramon Bragós and doctoral student Tomàs Garcia, who are attached to the Biomedical Engineering Research Centre (CREB), in collaboration with a team at the University of Barcelona (UB).

The UPC is also contributing to major advances in the development of medical devices and diagnostic imaging. The UPC’s Institute of Industrial and Control Engineering (IOC) and the Pulmonology Research Group of Bellvitge Hospital’s Institute for Biomedical Research have developed a virtual bronchoscopy system that improves the diagnosis of lung cancer. The technology provides doctors with information that enables them to decide with more confidence whether an actual bronchoscopy is necessary or not. This helps minimise risk and discomfort for patients.

The system is based on images provided by a virtual bronchoscopy using 2D computed tomography images. The novel feature of the system is that it takes into account the geometry and kinematic constraints of the bronchoscope.

The device is designed so that a pulmonologist can virtually navigate through a patient’s airways and simulate the movements that will later be executed when a flexible bronchoscope is used to perform the examination. It is a useful tool that facilitates “very realistic planning of the most feasible path from the trachea to peripheral pulmonary lesions,” says Jan Rosell, the researcher who carried out the project together with Paolo Cabras and Alexander Pérez, who also work with the IOC. “Doctors can also use the device to determine whether the end of the bronchoscope will reach a lesion, or, if not, how close it can be manoeuvred and what technique will need to be used to obtain a biopsy sample,” Rosell adds.

In addition to pursuing advances in diagnostic imaging, molecular biology and telemedicine, UPC researchers are also doing innovative work in another area of interest: metabolomics, the scientific study of chemical processes involving metabolites. It is in this field that another CREB team has patented an innovative software tool. The advanced program, based on a new algorithm, helps medical professionals make more accurate, automated predictions in disease diagnosis and drug screening.

Developed by Àlex Perera and Francesc Fernández in collaboration with researchers with the University of Barcelona’s Department of Nutrition and Food Science, the tool improves detection of biomarkers, the biological markers used to detect diseases.

Another advantage of the software is that it reduces prediction error in metabolomic analysis and testing (used to examine the small organic molecules in biological systems). Metabolomic analyses are based on biological samples of urine or blood, nuclear magnetic resonance (NMR) techniques, and mass spectrometry (LC/MS). Making predictions in this area is complex because it requires analysis of extensive data obtained from individual samples, but it is of vital importance in evaluating the effectiveness of new drugs, for example.

New test for tuberculosis

Tuberculosis is one of the diseases that accounts for the most morbidity and mortality worldwide. Despite this, there are still a lot of unanswered questions about the disease and many scientific challenges remain to be tackled. Daniel López Codina and Clara Prats of the UPC’s Discrete Modelling and Simulation of Biological Systems group have carried out research in this field in collaboration with a team at the Experimental Tuberculosis Unit of the Germans Trias i Pujol Health Sciences Research Institute Foundation.

The two teams have patented a new method that offers a fast, easy and reliable way to determine the virulence (ability to produce disease) of Koch’s bacillus. The technique allows specialists to make more accurate diagnoses.

López Codina’s team observed the tuberculosis bacillus (Mycobacterium tuberculosis) in an in vitro culture and looked at the way it grows by forming clumps. Given the difficulty of applying conventional microbiological methods with this type of culture, the researchers used an alternative approach: microscopy and analysis with image processing techniques. “This is the first time we’ve been able to use a culture to observe two different strains of the bacterial parasite and the existence of a correlation between the characteristic clumping pattern and the virulence of the disease,” said the researcher.

The results have created a new business opportunity for companies involved in biomedical imaging and diagnostic testing.

Projects like these highlight the huge potential of engineering and medicine to continue delivering solutions that improve people’s quality of life.

Source : http://www.news-medical.net/news/20121123/UPC-teams-conduct-research-in-biomedical-engineering-to-improve-peoples-health.aspx

Full story

Panasonic Develops A Novel Way to Sequence SNPs

Panasonic Develops A Novel Way to Sequence SNPs

Panasonic Develops A Novel Way to Sequence SNPs

Osaka, Japan – Panasonic, the leading brand by which Matsushita Electric Industrial Co., Ltd. is known, in collaboration with Professor Naoki Sugimoto of Konan University, has developed a technology for electrically identifying single nucleotide polymorphisms, or SNPs[1] (sequence variations in DNA[2]). This world-first technology* provides economical and accurate identification of SNPs by measuring electrical current without attaching DNA to electrodes.

This technology makes it possible to predict individuals’ responses to drugs and their risk of developing disease. In the future, this technology is expected to enable hospitals or clinics to provide patients with treatments and drugs tailored to their individual physical characteristics. With this new technology, Panasonic is contributing to a more personalized medical treatment based on the patient’s DNA sequence.

Conventional electrical SNP identification technologies, which use DNA-DNA hybridization reactions to identify the differences in DNA sequences, are not only inaccurate, but also require specialized and expensive electrodes to which artificial DNA needs to be attached.

Against this backdrop, Panasonic has achieved accurate SNP identification technology through the following new technologies:

Designing a base sequence of artificial DNA, for the DNA replication reaction, that depends strongly on the differences in DNA sequences.

Developing an electrical detection technology for phosphate compounds[3] released during DNA replication using the electrical current generated by enzyme reactions.

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Technological Features

Accurate SNP identification using a DNA replication reaction that depends strongly on differences in DNA sequences

To identify SNPs by using a replication reaction for the target DNA an artificially synthesized piece of single-strand short DNA consisting of approximately 20 bases is prepared. The base sequence of this artificial DNA is designed to correspond to that of the target DNA. If the SNP of the target DNA is complementary to the SNP-corresponding nucleotide of the artificial DNA, the replication reaction of the artificial DNA occurs; however, the replication reaction does not occur if the SNP is not complementary. A SNP can be identified accurately by using this reaction.

The world’s first electrical SNP identification without attaching artificial DNA to an electrode

Conventional electrical SNP identification requires specialized and expensive electrodes, on which artificial testing DNA is attached. However, there are still problems in controlling the quantity to be secured and the variation in the result. Another approach is an optical identification method using fluorescent dyes without attaching DNA to an electrode, but this requires large and expensive equipment along with an optics system.

Panasonic’s improved new method can identify SNPs electrically with artificial DNA dissolved in a solution instead of being attached to the electrode.

Details of the Method

Technology for designing a base sequence of artificial DNA for a DNA replication reaction that depends strongly on the differences in DNA sequences

Panasonic discovered that when artificial DNA is prepared with a base sequence that is non-complementary to the second and third bases from the SNP of the target DNA, a replication reaction from the artificial DNA occurs if the SNP is complementary to the SNP-corresponding nucleotide; however, no replication reaction occurs if it is not complementary.

Electrical detection technology for phosphate compounds released during DNA replication as a ‘current value’ generated by enzyme reactions

During DNA replication, a single molecule of a phosphate compound called pyrophoric acid is released as a reaction byproduct every time a base is replicated. Panasonic has developed a new enzyme reaction system for converting the quantity of pyrophoric acid into an electrical current. By using three types of enzymes, the quantity of pyrophoric acid is converted to an equivalent quantity of potassium ferrocyanide, an electron mediator, making measurement of the oxidation current possible, a world-first achievement.

Image of SNP Sensor Chip

Patents: 24 in Japan, 22 overseas (including those pending)

 

[1] SNP (Single nucleotide polymorphism)

Of the individual differences seen in the DNA sequence, the condition where one base pair is different from the others, as well as its position, is called a SNP. The three billion base pairs that make up human DNA are thought to contain more than a million types of SNPs. This diversity of base pairs is believed to generate individual differences in each person’s reaction to drugs, development of diseases, and other health factors.

[2] DNA (Deoxyribonucleic acid)

All living organisms need to synthesize proteins, and the blueprint for this is DNA. The sequence of the four types of base (Adenine [A], Guanine [G], Cytosine [C], and Thymine [T]) controls the types of protein synthesized and their quantity. Two strands of DNA chains form a double helix. In this structure, ‘A’ specifically forms a base pair with ‘T,’ and ‘G’ with ‘C.’ (A combination of bases that can form a base pair is called complementary, and other combinations that cannot form a base pair are called non-complementary). One of the two strands can make a copy of the original DNA to transmit genetic information.

[3] Phosphate compounds

As a byproduct of the DNA replication process, one molecule of a phosphate compound, called pyrophoric acid, is released every time one base is replicated.

* As of August 20, 2008

About Panasonic

Best known for its Panasonic brand name, Matsushita Electric Industrial Co., Ltd. is a worldwide leader in the development and manufacture of electronic products for a wide range of consumer, business, and industrial needs. Based in Osaka, Japan, the company recorded consolidated net sales of 9.07 trillion yen (approx. US$90.52 billion) for the year ended March 31, 2008. The company’s shares are listed on the Tokyo, Osaka, Nagoya and New York (NYSE:MC) stock exchanges. For more information on the company and the Panasonic brand, visit the company’s website at http://panasonic.net/.

Matsushita of Japan, commonly known as Panasonic, has developed a technique to electronically identify single nucleotide polymorphisms in a DNA solution without direct contact between the molecules and electrodes. Knowledge of one’s DNA variance (in the form of nucleotide polymorphisms) can point to differences in individual response to specific pharmaceuticals, as well as risks of having a certain disease.

Details of the technology:

To identify SNPs by using a replication reaction for the target DNA an artificially synthesized piece of single-strand short DNA consisting of approximately 20 bases is prepared. The base sequence of this artificial DNA is designed to correspond to that of the target DNA. If the SNP of the target DNA is complementary to the SNP-corresponding nucleotide of the artificial DNA, the replication reaction of the artificial DNA occurs; however, the replication reaction does not occur if the SNP is not complementary. A SNP can be identified accurately by using this reaction.

Conventional electrical SNP identification requires specialized and expensive electrodes, on which artificial testing DNA is attached. However, there are still problems in controlling the quantity to be secured and the variation in the result. Another approach is an optical identification method using fluorescent dyes without attaching DNA to an electrode, but this requires large and expensive equipment along with an optics system.

Panasonic’s improved new method can identify SNPs electrically with artificial DNA dissolved in a solution instead of being attached to the electrode.

Panasonic discovered that when artificial DNA is prepared with a base sequence that is non-complementary to the second and third bases from the SNP of the target DNA, a replication reaction from the artificial DNA occurs if the SNP is complementary to the SNP-corresponding nucleotide; however, no replication reaction occurs if it is not complementary.

During DNA replication, a single molecule of a phosphate compound called pyrophoric acid is released as a reaction byproduct every time a base is replicated. Panasonic has developed a new enzyme reaction system for converting the quantity of pyrophoric acid into an electrical current. By using three types of enzymes, the quantity of pyrophoric acid is converted to an equivalent quantity of potassium ferrocyanide, an electron mediator, making measurement of the oxidation current possible, a world-first achievement.

Source : http://panasonic.co.jp/corp/news/official.data/data.dir/en080819-2/en080819-2.html

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Abacus Diagnostica’s GenomEra C. difficile assay receives CE mark

Abacus Diagnostica’s GenomEra C. difficile assay receives CE mark

Abacus Diagnostica Ltd announced that it has received the CE mark for the GenomEra™ C. difficile assay for the detection of toxin-producing Clostridium difficile directly from stool samples. C. difficile is the leading cause of infectious nosocomial diarrhea in Europe and North America.

Rapid changes in the epidemiology and increasing incidence have taken the focus on methods to diagnose toxigenic C. difficile faster and more efficiently. Accurate and rapid diagnosis of C. difficile infection is crucial for patient care but also for preventing transmission and reducing the overall disease burden.

The GenomEra C. difficile assay is the newest test of Abacus Diagnostica’s expanding line of easy-to-use and cost-efficient molecular diagnostics products. The C. difficile test is extremely simple to perform directly from stool specimen through a straightforward sample preparation process without extraction, heating or centrifugation steps. The assay is built on rapid and reliable target amplification and end-point detection technology that allows for high quality results in 50 minutes.

“Abacus is committed to providing a continuum of diagnostic products for key infectious diseases to meet the needs of various laboratory settings,” said Tom Palenius, CEO of Abacus Diagnostica. “Along with our earlier CE marked MRSA/SA assays this PCR test will help to meet the testing and resource challenges of many different types of labs. During next year we will continue to expand our molecular test offering for critical blood stream and perinatal infections.”

Source : http://www.news-medical.net/news/20121119/Abacus-Diagnosticae28099s-GenomEra-C-difficile-assay-receives-CE-mark.aspx

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DNA Sequencing Moving Into The Clinic; Used to Diagnose Mitochondrial Disease in 42 Children

DNA Sequencing Moving Into The Clinic; Used to Diagnose Mitochondrial Disease in 42 Children

DNA sequencing has identified difficult-to-diagnose diseases in humans – the first time the technology has been used in a clinic.

The technique, which decodes thousands of genes simultaneously, has been used in laboratories to uncover genes related to diseases since 2009.

Now it has successfully moved to the clinic, where patients do not know what is wrong with them and may not know their family history of disease, and clinicians have few clues about which genes might be causing the problem.

Mitochondrial diseases, which affect the way the body produces energy, are notoriously difficult to diagnose. Found in at least one in every 5000 people, the diseases often involve many genes, and symptoms vary across organs. For example, common manifestations can include blindness, seizures, slow digestion and muscle pain.

Currently, diagnosing such disorders can take months or even years, and involves an invasive muscle biopsy. DNA sequencing technology may help to speed things up.

Diagnostic data

Elena Tucker and colleagues from the Murdoch Childrens Research Institute in Sydney, Australia, along with Vamsi Mootha from Harvard Medical School, sequenced the genomes of 42 children who had traits that suggested they carry a mitochondrial disorder. To work out exactly which disorder each child carries, the team looked both at the DNA in their mitochondria and at the 100 or so genes within their nuclear DNA that have already been linked to mitochondrial diseases. They also looked at a further 1000 nuclear genes that play a part in mitochondrial biology.

To distinguish between harmless genetic variations and those that might cause a disease, the team compared the patients’ genomes with databases of genetic variation recorded in the general population.

Ten of the children had mutations in genes previously linked to mitochondrial diseases, and so could be given a precise diagnosis. Mutations not previously associated with any disease were found in another 13 children. Tucker says that these patients can expect a full diagnosis once studies confirm the function of these genes.

“We are quite excited,” says Tucker. “Most of these diagnoses were in children whose [illnesses] could not easily be diagnosed using traditional methods.”

Needle in a haystack

Michael Ryan, a biochemist at La Trobe University in Melbourne, Australia, who was not involved in the work, says the diagnosis rate “will improve” within the next couple of years as the list of genes known to be linked to mitochondrial diseases grows, and it becomes clearer how mutations combine to cause diseases.

“It’s a fantastic study,” says Matthew McKenzie at Monash University in Melbourne. Finding genetic mutations in mitochondrial patients is “like searching for a needle in a haystack”, he says. “I think it was a very good result to transfer to a clinical setting.”

Advances in next-generation sequencing (NGS) promise to facilitate diagnosis of inherited disorders. Although in research settings NGS has pinpointed causal alleles using segregation in large families, the key challenge for clinical diagnosis is application to single individuals. To explore its diagnostic use, we performed targeted NGS in 42 unrelated infants with clinical and biochemical evidence of mitochondrial oxidative phosphorylation disease. These devastating mitochondrial disorders are characterized by phenotypic and genetic heterogeneity, with more than 100 causal genes identified to date. We performed “MitoExome” sequencing of the mitochondrial DNA (mtDNA) and exons of ~1000 nuclear genes encoding mitochondrial proteins and prioritized rare mutations predicted to disrupt function. Because patients and healthy control individuals harbored a comparable number of such heterozygous alleles, we could not prioritize dominant-acting genes. However, patients showed a fivefold enrichment of genes with two such mutations that could underlie recessive disease. In total, 23 of 42 (55%) patients harbored such recessive genes or pathogenic mtDNA variants. Firm diagnoses were enabled in 10 patients (24%) who had mutations in genes previously linked to disease. Thirteen patients (31%) had mutations in nuclear genes not previously linked to disease. The pathogenicity of two such genes, NDUFB3 and AGK, was supported by complementation studies and evidence from multiple patients, respectively. The results underscore the potential and challenges of deploying NGS in clinical settings.

The first thing Debbie Jorde noticed about her newborn daughter was that her arms were bent at unnatural angles. She had other problems, too: a cleft palate, eight fingers, eight toes and no lower eyelids. She would eventually be diagnosed with Miller syndrome, a disease so rare that doctors have long assumed that each case arises through spontaneous mutation, rather than being passed down through families. Doctors told Jorde that her chances of having a second child with the syndrome were less than one in a million.

They were wrong. Jorde’s son, born three years after his sister, had the same features. Lynn Jorde, Debbie’s current husband and a geneticist at the University of Utah in Salt Lake City, still cringes when Debbie recounts what the doctors had told her. “The right answer for that situation is that there have been so few cases that we really can’t predict the risk,” he says.

Thanks to next-generation genome sequencing, Debbie and her children now know the family’s genetic risks. Lynn and his collaborators had been talking about sequencing the genomes of an entire ‘nuclear’ family affected by a genetic disease, both to identify the mutation responsible and to investigate how genes are inherited in unprecedented detail. Debbie, her former husband and her now-adult children, Heather and Logan Madsen, were happy to be take part, and in 2009 became the first family in the world to have their genomes fully sequenced1.

Over the course of six months, the research team cross-compared the whopping amount of DNA data from the four genomes. With the help of a parallel sequencing effort that included others with Miller syndrome2, the researchers identified the gene involved, called DHODH, which encodes a protein involved in the synthesis of nucleotides. The disease, it turns out, is recessive. In this case, both parents carried a single mutated copy of the gene, so their chance of having a child with the syndrome was actually one in four. The analyses also revealed that the children had a second recessive genetic disorder, primary ciliary dyskinesia, which affects lung development. Before that discovery, says Debbie, “We never knew why they kept getting pneumonia.”

Click for larger image.

Families like Debbie Jorde’s are part of a small but growing vanguard of people, mostly with rare diseases and cancers, whose genomes have been sequenced to help diagnose or understand their condition. Although knowing the sequence didn’t alter treatments for Heather and Logan, some individuals are being sequenced with that intent. A boy in Wisconsin was given a risky but life-saving bone-marrow transplant last year on the basis of a partial genome sequence3; a woman with leukaemia was spared a similar procedure after her genome was sequenced4; and genome sequencing was used to refine the therapy given to twins with a rare disorder (see ’6 billion to one’)5.

Most of those involved so far have been lucky enough to know the right people — researchers with an interest in clinical genetics — or determined enough to seek them out, and many, such as Debbie Jorde’s family, were taking part in research projects. But now, with genome sequencing becoming much cheaper and faster, clinical programmes are starting up around the world that will routinely analyse genomes for those who might benefit from the information. Illumina, which is based in San Diego, California, and provided the sequencing machines for many of the programmes, offers whole-genome sequencing for as little as US$7,500 for people with life-threatening disease, and for $10,000 for people with cancers that require the sequencing of both tumour and non-cancer cells.

As prices fall further, some say that prescribing a genome sequence or analysis will become akin to requesting a magnetic resonance imaging (MRI) scan. “It’s just like any other test in medicine. There’s nothing remotely special about it,” says David Bick, a clinical geneticist at the Medical College of Wisconsin in Milwaukee. But, he adds, “people will cry and scream and yell about that statement”. That’s true: unlike the results of most medical tests, a genome sequence provides a vast amount of difficult-to-interpret data, not all of which will be necessary for diagnosing or treating the patient’s condition and which could provide unwanted clues to future health risks. The few success stories published so far also suggest that wringing information from the human genome and counselling patients and their families adequately may be too big a burden for medical systems that are already stretched to their limits. “You can’t immediately jump from those few profound but limited stories and think that you can reduce this to practice for clinical care,” says Eric Green, director of the National Human Genome Research Institute (NHGRI) in Bethesda, Maryland. Still, from the pioneering cases, much can be learned.

Rare births

Take Nicholas Volker. From the time he was born, an undiagnosed condition ravaged his intestines, sometimes causing fistulae: holes that ran from his gut through to the outside of his body, leaking faeces and requiring surgery. By the time he turned three, Volker had been in an operating room more than 100 times. Doctors hypothesized that he had an immune deficiency and that a bone-marrow transplant might correct the problem. But a number of tests, including the sequencing of several genes, were inconclusive. After intense deliberation, a team at the Medical College of Wisconsin was cleared to sequence Volker’s exome, the 1–2% of the genome that codes for proteins and key regulatory RNA molecules.

Using computational tools, the team combed Volker’s DNA for sequences that vary from person to person. They compared these with known variants in the general population, with variants associated with diseases and with related sequences in other species, looking for a mutation that might have caused the problem, says David Dimmock, a clinical geneticist at the college. It took, “basically one person staring at a computer for three and a half months”, he says, but eventually they identified a mutation on the X chromosome in a gene called X-linked inhibitor of Apoptosis, or XIAP (ref. 3). A deficiency of the protein encoded by this gene is known to put patients at high risk for a deadly immune-cell disorder, and a bone-marrow transplant suddenly became imperative. More than a year later, Dimmock says, Volker is doing well.

What started as an experiment has become a programme at Wisconsin, where Dimmock, Bick and their colleagues now aim to provide comprehensive whole-genome sequencing for patients. The team is focusing on people with rare disorders that are thought to involve a genetic defect, and in whom identifying that defect is likely to inform the course of treatment.

Bick says that of 48 patients evaluated for the programme, 17 have been accepted, and their families have gone through six hours or more of genetic counselling before sequencing. Insurance companies have agreed to foot the bill for at least two of the cases. Their rationale is straightforward, says Tina Hambuch, a senior scientist at Illumina’s clinical services laboratory, which has been doing the sequencing for this programme. A full genome sequence can be less expensive than a series of single genetic tests, and might clarify whether a costly treatment is required. “There are cases where it’s cost effective,” Hambuch says.

Genome factories

Other institutions are following suit. In the United Kingdom, the Wellcome Trust Centre for Human Genetics at the University of Oxford has made plans with Illumina to sequence 500 genomes from people — some from the same family — with a wide range of conditions. The Undiagnosed Diseases Program at the National Institutes of Health in Bethesda has been running a sequencing programme since 2008. It has sequenced more than 140 exomes and 5 genomes in its attempts to find the molecular underpinnings of diseases that have eluded diagnosis. The programme was so overwhelmed by interest that it temporarily stopped accepting applications a few months ago.

Green says that “now is the time to push the accelerator”. Clinical geneticists often talk about tackling Mendelian disorders: diseases thought to involve a single gene and that roughly obey the rules of inheritance drawn up by Gregor Mendel in the nineteenth century. These conditions may account for as many as 20% of paediatric hospitalizations worldwide and a large share of health-care costs. Yet their genetic basis is often unknown. The compendium of such conditions, called Online Mendelian Inheritance in Man (OMIM), currently contains just under 7,000 disorders, about half of which have been assigned a molecular cause. This autumn, Green says, the NHGRI will announce the winners of its Mendelian Disorders Genome Centers grants, which will fund sequencing centres looking for causes of the rest.

Still, many researchers worry that it will be difficult to make clinical use of most genomes. At the Undiagnosed Diseases Program, the misses have certainly outnumbered the hits so far. “I think we’ve learned a lot about how hard evaluating an exome is,” says Thomas Markello, from the medical-genetics branch of the NHGRI. “I’m most concerned that people don’t recognize that what’s been published to date are the success stories.”

“We’ve learned a lot about how hard evaluating an exome is.”

Many researchers say that genome sequencing could be used in diagnosis and therapy of cancer more easily than in rare diseases. Clinicians are already doing sophisticated analyses of some tumours in order to tailor therapies to the patient’s genetic characteristics; a genome sequence provides even more molecular detail. For example, an individual’s cancer genome sometimes reveals defects in a pathway that might point to use of a known drug, but were not apparent from standard tests.

In 2007, a 78-year-old man in Canada with a rare tongue cancer that had spread throughout his body was being treated at the British Columbia Cancer Agency in Vancouver. There was no approved treatment for his type of cancer and — being what doctors described as a “savvy sort” — he and his clinician convinced scientists at the agency to sequence the cancer’s genome. The scientists also analysed its transcriptome, revealing both the sequence and the amount of RNA that the tumour was producing. The team then compared these data with those for other cancers and for the patient’s normal cells.

The researchers homed in on RET, a gene known to promote cancer, which was duplicated in the tumour genome and churning out RNA. Several drugs are known to inhibit the protein encoded by this gene. Marco Marra, director of the cancer agency’s genome-sciences centre, says that “after much agonizing and hand-wringing”, the clinical team prioritized these drugs and tried the top one, sunitinib. The cancer stabilized for several months on this and a second treatment, but eventually started to spread again. An analysis of the recurring tumours showed that different cancer-promoting pathways had been activated6, making the tumours resistant to the first drug, but possibly responsive to others. Unfortunately, by then it was too late to do more: the man died.

Unstoppable train

Marra’s group is now setting up a project to better diagnose subtypes of another cancer, acute myelogenous leukaemia, using transcriptome and other sequencing methods. Partly inspired by Marra’s efforts, Elaine Mardis, a geneticist at Washington University in St Louis, Missouri, and her collaborators have used genome sequencing to try to help a handful of people with cancer, including the woman with leukaemia4. The woman had been treated and had gone into remission, but standard tests were unable show conclusively whether she had acute promyelocytic leukaemia (APL) — which generally has a good outcome with standard therapy — or a type of leukaemia that would require aggressive follow-up treatment, such as a bone-marrow transplantation. Over about seven weeks, the team sequenced the cancer’s genome and found a gene fusion that was consistent with APL. Mardis is enthusiastic about the approach, but notes its limitations. “It’s another piece of evidence,” she says. “It’s not going to be the only thing that you’re looking at when going to a patient diagnosis.”

Moving whole-genome sequencing from research to clinic is beset with challenges. Unlike in research, DNA sequencing that is intended to inform a diagnosis must be done in accredited laboratories, such as those used by Illumina. The institutional review boards that oversee research in humans have not reached a consensus on whether approval is needed for clinical genome sequencing; and the US Food and Drug Administration is yet to work out how to regulate the coming wave of clinical sequencing.

Many researchers and clinicians worry that health systems don’t have enough people well versed in genomics or bioinformatics to interpret the flood of data. What’s more, say experts, function and disease information for the human genome is scattered across scientific articles and databases that are hard to troll through and aren’t always correct. Sequence analysis is where most costs now lie. Hambuch says that for the few research projects on which Illumina has collaborated, just identifying all the variants in a genome has taken two to three weeks. “That’s a lot of effort from high-skill people,” she says.

The information could also overwhelm patients. Medical geneticists and ethicists have long worried about finding genetic pointers to disease risks that are unrelated to the illness being treated. With a full genome sequence, the likelihood of such incidental findings shoots up. The situation is particularly tricky for young patients. Do parents have the right to decide for them what information is revealed? This is where many of those hours of genetic counselling are spent, says Bick.

For these reasons, Stephen Kingsmore at Children’s Mercy Hospital in Kansas City, Missouri, argues that clinical sequencing should be limited in scope. He advocates sequencing just what he calls the Mendelianome, the genetic regions known to be involved in inherited diseases. “Ethically, legally, socially that’s going to be more acceptable,” he says. His group is developing methods that use a panel of mutations associated with just over 600 recessive diseases for such screening. Doing much more than this, he says, puts research goals ahead of the patient.

But some geneticists think that the train is unstoppable. “Once you demonstrate how informative this technology is, I think this is going to be widely adopted,” says Hakon Hakonarson, who is starting a programme for clinical assessment of genomes at the Children’s Hospital of Philadelphia in Pennsylvania.

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The members of Debbie Jorde’s family still ponder what their genome sequences have meant for them. Although the sequences didn’t alter treatment, if they had known about the lung problem earlier it might have prevented a dangerous procedure that both Heather and Logan underwent to reduce the recurrence of pneumonia.

Still, Lynn Jorde thinks that more successes are on the way for genomes in clinical care. “I’d predict some spectacular applications.” But, he adds, “I’m a congenital optimist”.

DNA sequencing is slowly making the long anticipated move from the research lab into clinical use. After a few isolated reports last year where it was used to diagnose individual patients, now we see one of the first reports of it being used successfully on a larger scale, in 42 infants suspected of mitochondrial disease.

Mitochondria carry their own DNA but also rely on nuclear DNA for part of their functioning, and mutations in either of the two can cause malfunctioning of the mitochondria. Mitochondrial diseases may lead to a wide variety of symptoms and can be hard to diagnose, despite recent advances in genetic and biochemical tests.

All the infants in the study had clinical and biochemical evidence of mitochondrial oxidative phosphorylation disease. The researchers performed “MitoExome” sequencing of the mitochondrial DNA (mtDNA) and exons of approximately 1000 nuclear genes encoding mitochondrial proteins. Firm diagnoses were enabled in 10 patients (24%) who had mutations in genes previously linked to disease. Thirteen more patients (31%) had mutations in nuclear genes not previously linked to disease. These patients can expect a full diagnosis once studies confirm the function of these genes.

A few years ago, or maybe even one year ago, the cost of sequencing one individual’s DNA was too prohibitive to even think of using clinically. Now that the cost of sequencing one genome has come down to around $1000 and will probably come down even lower, genome sequencing will soon be cheaper than ordering more than two or three individual genetic tests. Although the current study only completely sequenced the mitochondrial DNA, it is a great example of how the technology can aid in diagnosis and even point to the existence of previously unknown mutations as origins of disease. We eagerly await the day that full-genome sequencing is just another test that is performed within the time of your doctor visit who directly receives and interprets the results.

Source : http://www.newscientist.com/article/dn21391-first-diagnosis-of-disease-by-dna-sequencing.html

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New England Biolabs launches new NEBNext Ultra kits at ASHG annual meeting

New England Biolabs launches new NEBNext Ultra kits at ASHG annual meeting

New England Biolabs, Inc. (NEB) launched their new NEBNext® “Ultra” kits at the recent American Society for Human Genetics (ASHG) annual meeting. These kits provide streamlined, low-input methods to prepare DNA and RNA libraries for Illumina® next generation sequencing. The supplied protocols and reagents are designed to maximize useful data from a broad range of samples, including those available in limited amounts.

The NEBNext Ultra DNA and Ultra RNA Library Kits produce high-yield libraries from 5 ng to 1 ug of input DNA, or as little as 10 ng of input RNA. The input RNA can be total RNA, purified mRNA or rRNA-depleted RNA.

Early access user Cynthia Hendrickson, Ph.D., at the HudsonAlpha Institute for Biotechnology, reports that in initial experiments, NEB’s new method “has allowed us to reduce our DNA inputs for exome enrichment from micrograms to a few hundred nanograms, while providing a simpler, faster protocol that minimizes the amount of hands-on time required.”

Another early access user, Momchilo Vuyisich, Ph.D., at the Los Alamos National Laboratory, says, “After testing several NGS library prep kits, we have found that the NEBNext Ultra DNA kit for Illumina has the best overall utility for resequencing and assembly of bacterial genomes. The kit offers an unmatched combination of ease-of-use, low cost, robustness and low input DNA requirements. It consistently produces excellent sequencing data.”

The new Ultra kits contain novel ligation reagents, as well as NEB’s NEBNext NGS-optimized formulation of Q5® High-Fidelity DNA polymerase, which provides ultra high-fidelity amplification and minimized GC bias.

NEB’s streamlined protocol for constructing DNA libraries with the Ultra kit requires only 15 minutes of hands-on-time, and is complete in 2.5 hours. The Ultra RNA workflow also incorporates a streamlined protocol; the workflow is complete in 4-5 hours, with only 30 minutes of hands-on time.

Source : http://www.news-medical.net/news/20121117/New-England-Biolabs-launches-new-NEBNext-Ultra-kits-at-ASHG-annual-meeting.aspx

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Courtagen, AGBL enter exclusive distribution agreement for clinical tests

Courtagen, AGBL enter exclusive distribution agreement for clinical tests

Courtagen Life Sciences Inc. announced today they have signed an exclusive strategic distribution agreement with the Alliance Global Group (AGBL) to help clinicians in emerging markets diagnose and manage a number of genetically based disorders. Courtagen’s test menu currently offers clinical tests for mitochondrial disorders (mtSEEK™ and nucSEEK™) and a newly released panel for epilepsy and seizure (epiSEEK™). Courtagen plans to add a number of key tests for other important neurological disorders.

“We are pleased to enter into this agreement with Courtagen”

“Building this relationship with AGBL is a critical component of Courtagen’s international business expansion,” said Brian McKernan, CEO, Courtagen Life Sciences, Inc.

Courtagen’s CLIA Laboratory genetic tests use next generation sequencing (NGS) technology, propriety bioinformatics platform, ZiPhyr™ and state of the art analytic capabilities to accurately sequence patient’s DNA and translate the sequence data into highly useful, clinically relevant information for physicians. Additionally, Courtagen’s protein diagnostics division offers decentralized testing of protein biomarker to simplify pre-clinical and clinical trial protocols.

“We are pleased to enter into this agreement with Courtagen,” said Dr. Tamer Degheidy, CEO of Alliance Global. “Courtagen’s portfolio of genetic tests will certainly have a positive impact on patients’ diagnosis and management in markets covered by our group.”

Source : http://www.news-medical.net/news/20121115/Courtagen-AGBL-enter-exclusive-distribution-agreement-for-clinical-tests.aspx

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DNA sequencing helps detect MRSA outbreaks in hospitals

DNA sequencing helps detect MRSA outbreaks in hospitals

This study used DNA sequencing to examine an outbreak of MRSA in a hospital, to uncover new cases and, as the study developed, to intervene in the outbreak to end it more quickly. Sequencing illuminated each person infected and described the transmission of MRSA between people coming to the hospital and within the hospital. This is believed to be the first time that sequencing has been used to close an infectious outbreak and will be published in Lancet Infectious Diseases.

For the first time, researchers have used DNA sequencing to help bring an infectious disease outbreak in a hospital to a close.

Researchers from the Wellcome Trust Sanger Institute, the University of Cambridge and Cambridge University Hospitals used advanced DNA sequencing technologies to confirm the presence of an ongoing outbreak of methicillin-resistant Staphylococcus aureus (MRSA) in a Special Care Baby Unit in real time. This assisted in stopping the outbreak earlier, saving possible harm to patients. This approach is much more accurate than current methods used to detect hospital outbreaks.

Using this technology, the team revealed that the outbreak had extended into the wider community, a conclusion that could not be reached with available methods. They also used sequencing to link the outbreak to an unsuspecting carrier, who was treated to eradicate MRSA.

“We are always seeking ways to improve our patient care and wanted to explore the role that the latest sequencing technologies could play in the control of infections in hospitals,” says Dr Nick Brown, author, consultant microbiologist at the Health Protection Agency and infection control doctor at Addenbrooke’s Hospital Cambridge. “Our aim is to prevent outbreaks, and in the event that they occur to identify these rapidly and accurately and bring them under control.

“What we have glimpsed through this pioneering study is a future in which new sequencing methods will help us to identify, manage and stop hospital outbreaks and deliver even better patient care.”

Over a six month period, the hospital infection control team used standard protocols to identify 12 patients who were carrying MRSA. However, this standard approach alone could not give enough information to confirm or refute whether or not an ongoing outbreak was actually taking place.

In this study, the researchers analysed MRSA isolates from these 12 patients with DNA sequencing technology and demonstrated clearly that all the MRSA bacteria were closely related and that this was an outbreak. They also revealed that the outbreak was more extensive than previously realised, finding that over twice as many people were carrying or were infected with the same outbreak strain. Many of these additional cases were people who had recent links to the hospital but were otherwise healthy and living in the community when they developed a MRSA infection.

While this sequencing study was underway, the infection control team identified a new case of MRSA carriage in the Special Care Baby Unit, which occurred 64 days after the last MRSA-positive patient had left the same unit. The team used advanced DNA sequencing to show in real time that this strain was also part of the outbreak, despite the lack of apparent links between this case and previous patients. This raised the possibility that an individual was unknowingly carrying and transmitting the outbreak MRSA strain.

The infection control team screened 154 healthcare workers for MRSA and found that one staff member was carrying MRSA. Using DNA sequencing, they confirmed that this MRSA strain was linked to the outbreak. This healthcare worker was quickly treated to eradicate their MRSA carriage and thus remove the risk of further spread.

“Our study highlights the power of advanced DNA sequencing used in real time to directly influence infection control procedures,” says Dr Julian Parkhill, lead author from the Wellcome Trust Sanger Institute. “There is a real health and cost burden from hospital outbreaks and significant benefits to be gained from their prevention and swift containment. This technology holds great promise for the quick and accurate identification of bacterial transmissions in our hospitals and could lead to a paradigm shift in how we manage infection control and practice.”

In this instance, DNA sequencing was a key step in bringing the outbreak to a close, saving possible harm to patients and potentially saving the hospital money.

“Our study indicates the considerable potential of sequencing for the rapid identification of MRSA outbreaks,” says Professor Sharon Peacock, lead author from the University of Cambridge and clinical specialist at the Health Protection Agency. “What we need before this can be introduced into routine care is automated tools that interpret sequence data and provide readily understandable information to healthcare workers. We are currently working on such a system.

“If we have a robust system of this type in operation when the outbreaks occur, we predict that we will be able to stop them after the first few cases, as we will rapidly find clear connections.”

In their next step, the team will study all MRSA carriers and infected patients over the next year in Addenbrooke’s Hospital and surrounding hospitals and the community to understand transmission events with the aim of improving infection management.

Sir Mark Walport, Director of the Wellcome Trust, says: “This is a dramatic demonstration that medical genomics is no longer a technology of the future – it is a technology of the here and now. By collaborating with NHS doctors, geneticists have shown that sequencing can have extremely important applications in healthcare today, halting an outbreak of a potentially deadly disease.”

source : http://www.news-medical.net/news/20121114/DNA-sequencing-helps-detect-MRSA-outbreaks-in-hospitals.aspx

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MRSA outbreak tracked and contained using DNA sequencing

MRSA outbreak tracked and contained using DNA sequencing

Scientists have used DNA sequencing for the first time to effectively track the spread of, and ultimately contain, an outbreak of methicillin-resistant Staphylococcus aureus (MRSA), according to new research published in The Lancet Infectious Diseases.

The technique allowed researchers to map and control an MRSA outbreak in a special care baby unit (SCBU) much more effectively than traditional infection control techniques alone would have allowed, leading to hopes that in future, the management of MRSA and other harmful bacterial infections could be vastly improved by the routine use of DNA sequencing technologies.

Researchers from the Wellcome Trust Sanger Institute, the University of Cambridge and Cambridge University Hospitals initially performed a type of DNA sequencing known as whole-genome sequencing on MRSA isolates taken from 12 babies known to have been carrying MRSA during a 6 month period in 2011.

When the MRSA infections first arose, an infection-control team working in the hospital suspected that the cases were linked, but this could not be proven using conventional methods to track and characterise outbreaks, nor was it clear how the infection was spreading and what its source might be. In an attempt to halt the spread of infection, the infection control team recommended standard measures of decolonisation treatment to eradicate MRSA from carriers and a deep clean of the ward where the infections had occurred.

When the scientists retrospectively performed DNA sequencing on these MRSA isolates, they were able to confirm that the MRSA strains were closely related, and that the MRSA cases observed were therefore part of an outbreak. Moreover, by widening their analysis to include samples from parents and visitors to GP’s surgeries, they were able to determine that the outbreak had spread into the community, infecting twice as many people as previously suspected.

While this retrospective analysis was taking place, the hospital infection-control team identified a new case of MRSA carriage in the special care baby unit, more than 2 months after the last MRSA-positive patient had left the unit and the ward had been deep-cleaned. The researchers used rapid DNA sequencing to show that the new case of MRSA was related to the earlier outbreak, leading them to hypothesise that a member of staff in the hospital might be unwittingly carrying the MRSA strain identified months earlier, allowing the same strain to infect another patient months after the initial outbreak and infection control measures.

As a result of this, 154 health care workers were screened for MRSA, and one member of staff was found to be carrying the same strain of MRSA linked to the outbreaks. The worker was then treated to eradicate their MRSA carriage, and the outbreak was contained.

According to Professor Julian Parkhill, lead author from the Wellcome Trust Sanger Institute in Cambridge, UK, “Routine use of DNA sequencing could have detected this MRSA outbreak 6 months earlier than standard techniques, and might well have prevented substantial illness and costs arising from MRSA transmission and subsequent infection. Whole-genome sequencing of MRSA could make an important contribution to infection-control investigation and practice, allowing quicker identification, tracking and isolation of outbreaks than is currently possible.”

This is the first study in which DNA sequencing has been used alongside conventional methods in real time, allowing scientists to directly compare the two and to understand how DNA sequencing might be effectively used alongside existing techniques in future.

“Before this technology can be used in routine clinical practice, we will require automated tools that interpret sequence data and provide information to healthcare workers and people without specialist sequencing knowledge” says Professor Sharon Peacock, lead author from the University of Cambridge, who adds that “we are currently working on such a system”.

Writing in a linked Comment, Dr Binh Diep, from the University of California, San Francisco, USA, states that, “The advent of high-throughput whole-genome sequencing has the potential to revolutionise outbreak investigations by providing a substantial advance in our ability to discriminate between different strains, compared with traditional molecular methods.”

source : http://www.news-medical.net/news/20121114/MRSA-outbreak-tracked-and-contained-using-DNA-sequencing.aspx

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Researchers sequence the DNA of 6,700 exomes of cystic fibrosis patients

Researchers sequence the DNA of 6,700 exomes of cystic fibrosis patients

A multi-institutional team of researchers has sequenced the DNA of 6,700 exomes, the portion of the genome that contains protein-coding genes, as part of the National Heart, Lung and Blood Institute (NHLBI)-funded Exome Sequencing Project, one of the largest medical sequencing studies ever undertaken.

Scientists participating in the project initially expected that individual rare variants would have a greater effect on over 80 heart, lung and blood related traits and diseases of high public health significance, said Suzanne M. Leal, Ph.D., professor and director, Center for Statistical Genetics in the Department of Molecular and Human Genetics of Baylor College of Medicine in Houston, TX.

The researchers found that many (1.1 million) of the 1.2 million coding variants that they identified in exome data from 4,420 European-Americans and 2,312 African-Americans occurred very infrequently in the population and often were only observed in a single individual, explained Dr. Leal, who presented the findings today at the American Society of Human Genetics 2012 meeting.

Dr. Leal added that most of the observed coding variants are population specific, occurring in either European or African Americans. “Of the identified variants, about 720,000 change the genetic code in a manner that could produce flawed proteins. Yet the role played by most of these variants in disease development has not been established,” she said.

The major goal of the project was to understand how variation in the exome affects heart, lung and blood related traits and diseases.

The study participants were selected from a sample of over 220,000 individuals who participated in another National Institute of Health (NIH) supported study that had collected extensive medical data on the participants. “Individuals were selected to have a disease endpoint of interest or an extreme trait value of public health importance,” said Dr. Leal.

By sequencing the exomes of 91 cystic fibrosis patients, Dr. Leal and her research colleagues discovered and replicated an association between variants in the DCTN4 gene and when a patient first develops a Pseudomonas aeruginosa airway infection.

The researchers were also able to replicate many known associations between individual DNA variants and traits, such as high blood levels of low-density lipoprotein, known as the ‘bad’ cholesterol, and C-reactive protein, which increases the body’s response to inflammation.

The majority of these findings are for variants that are common in the population, said Dr. Leal.

To detect associations with rare variants, analyses were performed by aggregating information from individual variants within a gene. This approach successfully detected an association with rare variants in the APOC3 gene that lowers triglyceride levels, an unhealthy type of fat in the blood, said Dr. Leal.

“In order to detect associations with rare variants, due to their modest effects, very large samples sizes are required. In many cases the data from the Exome Sequencing Project gave us leads that had to be evaluated using more study subjects. One mechanism for doing this was by genotyping additional samples using the exome chip, which contains approximately 240,000 coding variants. The Exome Sequencing Project played a very important role in the development of the exome chip, by being the largest contributor of data,” she added.

According to the NHLBI, exome sequencing is an efficient way to search for rare variants associated with complex traits. In contrast to previous genome wide association studies (GWAS), which concentrated on common variants scattered throughout the genome, exome sequencing has the potential to accelerate the search for unambiguous genetic links to disease by focusing attention on the protein coding portion of the genome

In the journal Science, Dr. Leal and her colleagues wrote that GWAS have substantially improved knowledge about common genetic variation, but have been generally uninformative about the patterns of rare variation within the protein coding regions of the genome.

“This is a very new field for which new methodology had to be developed. We learned many lessons in the quality control and analysis of exome data, as well as what types of results one would expect to see when analyzing rare variants. Additionally, the Exome Sequencing Project has been extremely valuable in obtaining a better understanding of population genomics and the history of man,” Dr. Leal said.

source : http://www.news-medical.net/news/20121107/Researchers-sequence-the-DNA-of-6700-exomes-of-cystic-fibrosis-patients.aspx

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