Bioengineering Efficient Antibiotic Biosynthesis In E. Coli

The pathways underlying the production of antibiotics are now quite well known. For example, the antibacterial activity of erythromycin, an important polyketide antibiotic precursor, requires the transfer of two unusual sugars called mycarose and desosamine (both glycosyl groups) onto the nonsugar part of the glycoside molecule (macrocyclic aglycone).

In a new study published online this week in the open-access journal PLoS Biology, Ho Young Lee and Chaitan Khosla demonstrate how they used bioassay-guided evolution of this antibiotic pathway in Escherichia coli (E. coli) to identify more efficient antibiotic-producing mutants.

The authors reconstituted the biosynthetic pathways of both sugars in E. coli to yield the 6-deoxyerythromycin D antibiotic. By engineering a recombinant strain of E. coli that produces the bioactive macrolide 6-deoxyerythromycin D from propionate, they developed a fundamentally new tool for enhancing the efficiency of biosynthetic engineering of this class of antibiotics. Initially, this recombinant strain produced barely enough antibiotic activity to establish an activity-based screening assay. The authors therefore used the assay to screen for antibiotic overproducers. After three rounds of screening, they were able to identify E. coli cells that overproduced the 6-deoxyerythromycin D antibiotic with significant modifications in the mycarose biosynthetic pathway. They used the same activity-based screening system to evolve E. coli mutants capable of more efficient precursor-directed biosynthesis. As the first example of bioassay-guided evolution of an antibiotic pathway in E. coli, these results open the door for harnessing the power of genetics for mechanistic investigations into polyketide synthases and also for biosynthetic engineering.

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Citation: Lee HY, Khosla C (2007) Bioassay-guided evolution of glycosylated macrolide antibiotics in Escherichia coli. PLoS Biol 5(2): e45. doi:10.1371/journal.pbio.0050045.

CONTACT:
Chaitan Khosla
Stanford University
Stauffer III
381 North-South Mall
Stanford, CA 94305-5025

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Contact: Natalie Bouaravong
Public Library of Science

AIDS Memorial Quilt Should Be Displayed In Entirety To Raise Profile, Understanding Of HIV/AIDS, Letter To Editor Says

The Atlanta-based Names Project Foundation — which is involved in a lawsuit over the AIDS Memorial Quilt with Cleve Jones, who started the quilt in 1987 and served as its spokesperson for 15 years — should “let go” of the quilt and “assist in displaying it in its entirety” to raise the profile and understanding of HIV/AIDS in the U.S., Jerry Clark, founder and CEO of the not-for-profit We Care Minnesota, writes in a New York Times letter to the editor in response to a Jan. 31 Times article (Clark, New York Times, 2/4). Jones and NPF in December 2005 reached an agreement on the issue of returning a portion of the quilt to San Francisco. Under the agreement, Jones will receive 35 blocks of the quilt after he creates a San Francisco-based organization to oversee them. According to the agreement, Jones was required to establish a “501(c)(3) nonprofit organization that will have the name of San Francisco Bay Area Friends of the AIDS Memorial Quilt” by Dec. 31, 2006. The settlement stipulated that if the deadline was not met, the foundation would be “relieved of any obligation to supply blocks of the quilt to the nonprofit.” Jones said that he established a not-for-profit by the deadline through the San Francisco-based Tides Center, which oversees more than 200 projects. The dispute arises from the Tide Center’s Web site, which states that it is “legally and financially responsible for all Tides Center projects and activities” and that “[p]rojects are not separate entities or affiliated organizations — projects are Tides Center.” An attorney for NPF in January said Jones has not met the requirement (Kaiser Daily HIV/AIDS Report, 1/31). “If the quilt were displayed in its entirety, the country might just wake up” to the impact of HIV/AIDS, Clark writes, adding that increased funding for research and prevention efforts “would be a natural result of raising the profile and understanding of this terrible disease” (New York Times, 2/4).

“Reprinted with permission from http://www.kaisernetwork.org. You can view the entire Kaiser Daily Health Policy Report, search the archives, or sign up for email delivery at http://www.kaisernetwork.org/dailyreports/healthpolicy. The Kaiser Daily Health Policy Report is published for kaisernetwork.org, a free service of The Henry J. Kaiser Family Foundation . © 2005 Advisory Board Company and Kaiser Family Foundation. All rights reserved.

Human Skin Has Many More Types Of Bacteria Than Previously Thought

It appears that the skin, the largest organ in our body, is a kind of zoo and some of the inhabitants are quite novel, according to a new study. Researchers found evidence for 182 species of bacteria in skin samples. Eight percent were unknown species that had never before been described.

It is the first study to identify the composition of bacterial populations on the skin using a powerful molecular method. Not only were the bacteria more diverse than previously estimated, but some of them had not been found before, says Martin J. Blaser, M.D., Frederick King Professor and Chair of the Department of Medicine and Professor of Microbiology at NYU School of Medicine, one of the authors of the study.

“The skin is home to a virtual zoo of bacteria,” he says. This study is published February 5, 2007, in the online edition of the Proceedings of the National Academy of Sciences.

The researchers analyzed the bacteria on the forearms of six healthy subjects; three men and three women. “This is essentially the first molecular study of the skin,” says Dr. Blaser. The skin has been, he says, terra incognita, an unknown world that he and his colleagues have set out to understand much like explorers.

“There are probably fewer than ten labs in the U.S. looking at this question,” says Dr. Blaser. “It is very intensive work,” he adds. Zhan Gao, M.D., senior research scientist in Dr. Blaser’s lab, led the research, which took more than three years to complete.

Some of the bacteria on the skin appear to be more or less permanent residents; others are transient, according to the study.

This research is part of an emerging effort to study human microbial ecology. Dr. Blaser’s laboratory has previously examined the bacterial population in the stomach and the esophagus. “Many of the bacteria of the human body are still unknown,” he says. “We all live with bacteria all our lives and occasionally we smile, so they’re not that bad for us.”

The most numerous cells in our body are microbial – they outnumber our cells 10 to 1. The body has microbes native to the body, including the skin, and these populations change according to how we live, he says. “Ultimately what we want to do is compare disease and health,” says Dr. Blaser. Keeping bacterial populations in our body stable may be part of staying healthy, he says.

In the new study, the researchers took swabs from the inner right and left forearms of six individuals picking the region halfway between the wrist and the elbow for its convenience. “It’s not where they wash their hands,” explains Dr. Blaser. “And they don’t have to undress.” The researchers wanted to be able to compare two similar parts of the body. Because they also wanted to study change over time, they took swabs from four of the individuals 8 to10 months after the first test.

Roughly half, or 54.4%, of the bacteria identified in the samples represented the genera Propionibacteria, Corynebacteria, Staphylococcus and Streptococcus, which have long been considered more or less permanent residents in human skin.

The six individuals differed markedly in the overall composition of the bacterial populations on their skin. They only had four species of bacteria in common: Propionibacterium acnes, Corynebacterium tuberculostearicum, Streptococcus mitis, and Finegoldia AB109769. “This is a surprise,” says Dr. Gao. “But many things affecting the skin affect bacteria, such as the weather, exposure to light, and cosmetics use.”

Almost three-quarters, or 71.4%, of the total number of bacterial species were unique to individual subjects, suggesting that the skin surface is highly diversified in terms of the bacteria it harbors, according to the study.

Three bacterial species were only found in the male subjects: Propionibacterium granulosum, Corynebacterium singulare, and Corynebacterium appendixes. While the sample is too small to draw conclusions, the scientists believe that women and men may harbor some different bacterial species on their skin.

In each individual, the bacterial populations varied over time while revealing a core set of bacteria for each individual. “The predominant bacteria don’t change much,” says Dr. Gao. “But the more transient bacteria did change over time,” she says.

“What that suggests,” adds Dr. Blaser, “is that there is a scaffold of bacteria present in everybody’s skin. Some stay and others come and go.”

Finding the method

To obtain a sample Dr. Gao rubbed a swab on each individual’s forearms. “We didn’t tell them to be particularly clean, we just made sure they didn’t take antibiotics up to one month prior to the test,” Dr. Gao explains. She chose three men and three women to have a balance of genders. She set up a clean room so the samples didn’t risk contamination.

Traditionally, bacteria are cultured in the lab in petri dishes, which contain a medium to grow bacteria. But the method leads to inaccuracies, she explains, because only a fraction of bacteria in a sample grow in that medium. So the team used a powerful molecular method that involved extracting a subunit of genetic material called 16S ribosomal DNA from the samples. “It is kind of a common currency, it’s a conserved gene,” says Dr. Blaser. Another advantage is that there is a large database of 16S ribosomal DNA available to scientists.

The ambitious task for this study was to gather samples, prepare them, amplify the bacteria creating colonies of each single species of bacteria present in the skin samples. Then Dr. Gao used established tools – primers – to pick out the species-specific genetic regions in the bacteria. After sequencing those regions, the 16S ribosomal DNA (rDNA) in each colony, she consulted 16S rDNA databases to determine the bacterial species present in each sample. Many bacteria in the database only exist as sequences and have nether been named or extensively studied. Those are termed SLOTUs, or species-level taxonomic units.

Taxonomy and the study results

To distinguish organisms from one another, biologists group and categorize them. Species or SLOTUs are small categories. There are larger groupings such as genera and phyla. Humans, for example, belong to the phylum chordata, the genus Homo and the species Homo sapiens.

The molecular method used in this study revealed differences between the bacterial populations in individuals. Other methods had previously not shown those differences.

The team found a total of 182 species or SLOTUs and 91 genera of bacteria in the skin samples.

The samples yielded mainly three phyla of bacteria: Actinobacteria, Firmicutes, and Proteobacteria. Ninety-four point six percent of the bacteria were in these phyla. These phyla were found in all six tested individuals. When compared with earlier studies, the researchers found that these three phyla are also dominant in the esophagus and the stomach. In terms of bacterial species, however, the insides of the body, for example the stomach, and the exterior of the body, the skin, show vast differences in bacterial populations.

Skin condition can change markedly due to a variety of factors such as climate, diet, personal hygiene, and disease. But skin is never devoid of bacteria, particularly its more permanent residents. That is not bad news, after all, in healthy individuals these bacteria are not pathogens. “Without good bacteria, the body could not survive,” says Dr. Gao.

The next step for the research team is to look at diseased skin. “We plan to ask the question: Are the microbes in diseased skin, in certain diseases like psoriasis or eczema, different than the microbes in normal skin?” says Dr. Blaser.

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The study was funded by the National Institutes of Health (NIH), by a Senior Scholar Award from the Ellison Medical Foundation, and by the Diane Belfer Program in Human Microbial Ecology in Health. The authors of the study are Zhan Gao, M.D, Chi-hong Tseng, Ph.D., Zhiheng Pei, M.D., Ph.D, and Martin J. Blaser, M.D.

Contact: Jennifer Berman
New York University Medical Center and School of Medicine

Public Responses To The Recent Spinach Recall

Every year, the Food and Drug Administration issues dozens of food-related recalls, withdrawals and advisories. But few receive the attention that the advisory regarding E.coli-contaminated spinach received in September 2006. The broad scale of the resulting recall and related media attention provided a unique opportunity for researchers at the Rutgers Food Policy Institute (FPI) to study the U.S. food recall system. The results of this study were published on FPI’s web site, http://foodpolicyinstitute.org/.

To investigate the public’s reactions to this incident, a nationally representative sample of 1,200 Americans were interviewed by telephone from November 8 to 29, 2006. The results of the nationwide telephone survey describe the level of consumer awareness and knowledge of the recall and foodborne illness. The results also provide insight into consumer behavior during the recall and likely future behavior in response to the recall.

“We examined both the successes and of the failures of this particular recall,” said William Hallman, Director of the Food Policy Institute. “Our survey not only provides data to improve communications about future food recalls, but also enables us to explore how our systems might work in the case of intentional food contamination.”

The results of the survey show that the FDA’s main message to consumers warning that bagged fresh spinach had been contaminated and should not be eaten was heard by 87% of Americans. More than eight in ten (84%) of those who had heard about the recall said that they had also talked about it with others. In addition, the data clearly indicate that the majority of consumers did stop eating spinach because of the recall.

“As a result, the main public health goal of the recall was met,” said Hallman, “However, fewer Americans were aware of important details related to the recall. Many were confused about the types of spinach affected, where it was grown, the organism that caused the contamination, the symptoms of the resulting illness, and perhaps most significantly, whether or not the recall had ended.”

While nearly all (95%) of those who had heard about the recall knew that bagged fresh spinach had been recalled, only about two-thirds (68%) knew that loose fresh spinach was also part of the recall. However, they were confused about the safety of frozen and canned spinach during the recall, as only 57% and 71%, respectively, knew they were not affected by the recall.

Only half (52%) knew that the contaminated spinach had been grown in California, and only half (52%) could identify E. coli as the contaminant that made people ill. In addition, while 87% of Americans correctly recognized that abdominal cramps are a common symptom of E. coli infection, only about two-thirds (64%) of Americans correctly recognized the key symptom, bloody diarrhea. Instead, Americans are more likely to incorrectly associate the symptoms of nausea (88%) and vomiting (87%) with an E. coli infection. Moreover, though not generally associated with E. coli infections, more than three-quarters (77%) of Americans identified fever as a symptom, and nearly one-quarter (22%) reported that rashes were a symptom despite the fact that they are not commonly associated with any foodborne illness.

“Most Americans know little about the symptoms of foodborne illnesses,” said Hallman, “E. coli infections are no exception.” Although the recall caught the attention of the American public, not everyone followed the advice of the FDA. More than one-in-ten (13%) of those who ate spinach before the recall reported that they ate fresh spinach during the recall, and nearly three-quarters (74%) of them knew about the recall at the time.

Some Americans went to the other extreme, generalizing the warnings about spinach to other similar foods. Nearly one-fifth (18%) of those aware of the recall said they stopped buying other bagged produce because of the spinach recall. In addition, nearly half (48%) reported that the spinach recall caused them to wash their food more thoroughly.

“Clearly, the recall had a bigger effect on the public than just throwing away a few bags of spinach,” Hallman notes, “Consumers confidence in the safety of other produce seems to have been affected.”

While almost all Americans got the initial message that they should not eat fresh spinach, many fewer got the message that it is safe to eat it again. As of November 2006, many people were confused about the status of the recall. Thirteen percent believed that it was still in effect, and 18% said they did not know if it was still ongoing.

However, most spinach-eaters who knew about the recall said that they were already eating spinach again (44%) or may go back to eating spinach (47%), many within the next several months. Most Americans view their likelihood of getting sick from eating spinach as lower after the recall than during or even prior to the recall. Nonetheless, 5% of spinach-eaters who were aware of the recall said that they will never go back to eating spinach.

The authors of the study include Cara L. Cuite, Sarah C. Condry, Mary L. Nucci and William K. Hallman, all researchers at FPI. FPI is a research unit of Rutgers New Jersey Agricultural Experiment Station. The institute addresses important emerging food policy issues and supports public and private decision makers who shape aspects of the food system within which government, agriculture, industry and the consumer interact.

Contact: Michele Hujber
Rutgers, the State University of New Jersey

New Washington, D.C., HIV/AIDS Administration Director Pane Announces Initiatives To Fight City’s Epidemic

The director of Washington, D.C.’s Administration for HIV Policy and Programs, Gregg Pane, who replaced Marsha Martin in January, has announced plans to launch a series of “critical tasks” within the next 90 days aimed at addressing the district’s HIV/AIDS epidemic, the AP/Washington Examiner reports (AP/Washington Examiner, 2/4). District Mayor Adrian Fenty in January confirmed that he would not reappoint Martin as director of the city’s HIV/AIDS administration. Martin’s 16-month tenure as director of the HIV administration earned mixed reviews. Although she was lauded for increasing awareness about HIV/AIDS in the district, she was criticized for her method of collaborating and coordinating with the HIV/AIDS community (Kaiser Daily HIV/AIDS Report, 1/4). Pane’s HIV/AIDS initiatives include developing response and prevention plans, increasing condom distribution and enhancing HIV/AIDS case tracking, according to the AP/Examiner. Under the campaign, the administration by mid-February will distribute 250,000 condoms to 60 not-for-profits, and HIV rapid test kits will be distributed to physicians. The administration also is developing a comprehensive HIV prevention plan that will target young people, assess grant management and conduct oversight visits of the government’s service providers. In addition, Pane said that the administration plans to enter 1,200 backlogged HIV surveillance cases into city databases, which are needed to accurately record the extent of the epidemic and apply for federal grant money, the AP/Examiner reports. “I felt action was needed,” Pane said last week, adding, “We’re all saying it’s a crisis. Let’s do something. Shake it up and set some goals.” The DC Appleseed Center for Law and Justice contributed to planning the initiatives, Pane said. Appleseed Center Executive Director Walter Smith said, “My reaction is it’s a very good thing that Dr. Pane this quickly is trying to get organized and give this thing high priority, high visibility” (AP/Washington Examiner, 2/4).

“Reprinted with permission from http://www.kaisernetwork.org. You can view the entire Kaiser Daily Health Policy Report, search the archives, or sign up for email delivery at http://www.kaisernetwork.org/dailyreports/healthpolicy. The Kaiser Daily Health Policy Report is published for kaisernetwork.org, a free service of The Henry J. Kaiser Family Foundation . © 2005 Advisory Board Company and Kaiser Family Foundation. All rights reserved.

Texas AG Abbott Rules That State Law Permits Prison Officials To Mandate HIV Testing Among State Prisoners

Texas Attorney General Greg Abbott (R) on Thursday ruled that state law permits the Texas Board of Criminal Justice to mandate HIV testing among inmates upon entry to state prisons, the Houston Chronicle reports (Babineck, Houston Chronicle, 2/2). The state prison system in August 2006 had proposed a change to its HIV testing policy from saying new inmates “should be tested” upon entering prison to saying they “shall be tested” unless they opt out of testing. State Sen. Rodney Ellis (D) in August 2006 asked Abbott to rule whether current state law allows mandatory HIV testing for prisoners upon entry. Ellis — who with state Rep. Yvonne Davis (D) sponsored legislation approved last year that requires inmates to be tested for HIV before departing prison — said if Abbott ruled that current law does not allow mandated entry tests, he would sponsor a bill that would include such a requirement. About 80% of inmates have agreed to take an HIV test upon entering prison since the state began its testing program, and prison system statistics show more than 38,700 inmates received HIV tests in 2005. Of those, 372 tested HIV-positive. Texas law mandates that HIV test results are confidential and that HIV-positive inmates are not separated from HIV-negative inmates. Advocates for mandatory HIV testing upon entry into the prison system say it would help prison officials properly treat HIV-positive people, would provide more accurate data on the spread of the disease and could help officials estimate how many people are becoming HIV-positive in prison. July 2006 statistics show that of 154,000 prisoners in Texas, 2,627 are HIV-positive (Kaiser Daily HIV/AIDS Report, 8/24/06). Abbott’s opinion, which leaves the decision regarding mandatory testing up to the Texas Department of Criminal Justice, says that TBCJ is “authorized to adopt a rule or policy requiring mandatory” HIV testing among “incoming offenders in both the institutional division and the state jail division.” According to TDCJ spokesperson Jason Clark, there is no indication that the department is planning to make HIV testing among incoming prisoners mandatory anytime soon. Ellis did not comment on Abbott’s ruling, the Chronicle reports (Houston Chronicle, 2/2).

“Reprinted with permission from http://www.kaisernetwork.org. You can view the entire Kaiser Daily Health Policy Report, search the archives, or sign up for email delivery at http://www.kaisernetwork.org/dailyreports/healthpolicy. The Kaiser Daily Health Policy Report is published for kaisernetwork.org, a free service of The Henry J. Kaiser Family Foundation . © 2005 Advisory Board Company and Kaiser Family Foundation. All rights reserved.

Compounds Show Significant Promise Against Potential Bioweapon Toxins

Because of the high cost and limited applicability of currently available treatments, the newly identified compounds have the potential to fill the existing therapy gap and to provide protection against a bioterrorism attack using the toxin.

The study is being published the week of February 5 in an online edition of the Proceedings of the National Academy of Sciences.

“Our study is an important milestone in the fight against biological terrorism,” said Kim Janda, a Scripps Research scientist who led the study. “These small molecules are the first to show efficacy against this neurotoxin in animal models. Equally important, both have surprisingly simple structures, so their biological activity can be readily optimized. With their different modes of action, they could easily be developed as part of a potent ‘cocktail’ therapy.”

Janda said he expects to develop more small molecule candidates as potential botulism treatments.

Botulinum neurotoxins, which cause the disease botulism, are some of the most toxic substances known to scientists. One subtype, botulinum neurotoxin A, is a 100 billion times more potent than cyanide and relatively easy to produce, making it a potential biological weapon.

Using a multifaceted screening approach, Janda and his colleagues identified the two compounds and tested their efficacy in both cell-based assays and in mice exposed to the toxin.

One compound extended survival time by 36 percent (from 484 minutes to 659 minutes) a remarkable achievement considering its simple structure. Moreover, 16 percent of the animals treated with the second molecule survived with no obvious symptoms of botulism. No significant side effects were observed with either molecule.

Janda pointed out that the two compounds showed surprisingly little activity in cellular assays, suggesting that these standard cell-based screening methods may miss promising therapeutic candidates.

“Our study showed no correlation between cellular activity and in vivo efficacy,” Janda said, “which is highly unusual. Clearly, cell-based assays do not provide all the necessary information – animal-based studies are still an essential part of the discovery process. These findings validate our multidisciplinary screening approach to identify unrecognized chemical structures as potential treatments.”

While research efforts aimed at finding treatments for bioterrorism agents have increased dramatically since September 11, 2001, remarkably few have emerged. There are currently no small chemical molecules approved for treatment of botulism.

Botulism is a serious but extremely rare illness. There are seven related botulinum neurotoxins (A through G), although each acts differently and only four attack humans. The toxins kill through paralysis of the respiratory muscles. After attaching themselves to receptors on the neuronal surface – primarily muscle controlling motor neurons activated by acetylcholine, a neurotransmitter – the toxins block the release of acetylcholine proteins, inducing paralysis. Approximately 110 cases of botulism are reported each year in the United States.

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Other authors of the study, “An In Vitro And In Vivo Disconnect Uncovered Through High Throughput Identification Of Botulinum Neurotoxin A Antagonists,” are Lisa M. Eubanks, Mark S. Hixon, and Tobin J. Dickerson of The Scripps Research Institute, The Skaggs Institute for Chemical Biology, and The Worm Institute of Research and Medicine; Wei Jin, Sukwon Hong and Dale L. Boger of The Scripps Research Institute and The Skaggs Institute for Chemical Biology; Colin M. Clancy, Eric A. Johnson and William H. Tepp of the University of Wisconsin; Michael R. Baldwin and Joseph T. Barbieri of the Medical College of Wisconsin; Carl J. Malizio and Michael C. Goodenough of Metabiologics.

The study was supported by the National Institutes of Health, the National Institute of Allergy and Infectious Diseases and The Skaggs Institute for Chemical Biology.

About The Scripps Research Institute

The Scripps Research Institute is one of the world’s largest independent, non-profit biomedical research organizations, at the forefront of basic biomedical science that seeks to comprehend the most fundamental processes of life. Scripps Research is internationally recognized for its discoveries in immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune, cardiovascular, and infectious diseases, and synthetic vaccine development. Established in its current configuration in 1961, it employs approximately 3,000 scientists, postdoctoral fellows, scientific and other technicians, doctoral degree graduate students, and administrative and technical support personnel. Scripps Research is headquartered in La Jolla, California. It also includes Scripps Florida, whose researchers focus on basic biomedical science, drug discovery, and technology development. Currently operating from temporary facilities in Jupiter, Scripps Florida will move to its permanent campus in 2009.

Contact: Keith McKeown
Scripps Research Institute

Discovery Could Lead To Better Control Of Hemorrhagic Fever Viruses

Researchers report discovering the receptor through which a group of life-threatening hemorrhagic fever viruses enter and attack the body’s cells, and show that infection can be inhibited by blocking this receptor. The findings, to be published online by the journal Nature on February 7, give a clue to the high lethality of New World arenaviruses, suggest a way of reducing the severity of infection, and point the way toward a sorely needed treatment strategy.

The four viruses, known as the Machupo, Guanarito, Junin and Sabia viruses, cause Bolivian, Venezuelan, Argentine and Brazilian hemorrhagic fever, respectively, with mortality rates of about 30 percent. No vaccine is available, though a weakened form of Junin virus has been given to Argentinean farmers with some success. In addition to causing occasional disease outbreaks, mostly in poor, rural areas of South America, the viruses are of U.S. government interest because of their potential as bioterrorism agents. All four are classified as NIAID Category A Priority Pathogens and must be handled in Biosafety Level 4 containment facilities.

The researchers, led by Hyeryun Choe, PhD, of Children’s Hospital Boston’s Pulmonary Division, and Michael Farzan, PhD, of Harvard Medical School (HMS), first investigated the Machupo virus. To identify its cellular receptor, they made copies of the “spike” protein, used by the virus to gain entry into cells, and added it to cells from African green monkeys, known to be highly susceptible to Machupo virus infection. Later, they broke the cells open and isolated the spike protein and the cellular protein to which it had attached itself. Then, using a technique called mass spectrometry, they analyzed this attached cellular protein to determine its identity.

The receptor, identified in Choe’s lab by Jonathan Abraham, PhD, an MD-PhD student at HMS, turned out to be transferrin receptor 1 (TfR1), a well-known protein that is key in enabling cells to take up iron. Additional studies, performed in Farzan’s lab by HMS graduate student Sheli Radoshitzky, confirmed that TfR1 is also the receptor for the other three New World arenaviruses. (Abraham and Radoshitzky are both first authors on the study.) Expertise from Nancy Andrews, MD, PhD, an expert in iron metabolism at Children’s, sped up the work.

Although not all hemorrhagic fever viruses use TfR1 to enter the body’s cells, the discovery may help explain why these viruses wreak such havoc, damaging multiple organs and causing bleeding under the skin, in internal organs, and from orifices like the mouth, eyes or ears.

Because of TfR1’s essential function in transporting iron into cells, it is found on the surface of virtually every cell of the body. It is abundant on endothelial cells, which line blood vessels, a fact that may help account for the bleeding and organ damage caused by the viruses. TfR1 is also especially abundant on activated immune cells пїЅ” the very cells that mobilize to fight the viruses making them especially vulnerable to infection.

“This may help explain why mortality is so high,” says Choe, the study’s senior author.

Choe now hopes to translate these findings into treatments to contain natural or intentional outbreaks of New World hemorrhagic fever. Serendipitously, several anti-TfR1 antibodies have already been developed as anticancer therapeutics (cancer cells are also high in TfR1), and some have already been through clinical trials. Choe’s lab will test these antibodies, hoping to find one that inhibits virus entry without compromising TfR1’s essential function in cellular iron uptake.

“If some of these antibodies work, they could be used clinically fairly soon,” Choe says.

Coincidentally, Stephen Harrison, PhD, a structural biologist and Howard Hughes Medical Institute investigator at Children’s, had crystallized TfR1 and determined its 3-dimensional structure in 1999. Knowledge of TfR1’s structure will speed up the Choe lab’s efforts to pinpoint the parts of the molecule that are exploited by New World hemorrhagic fever viruses, which is necessary for the development of targeted antiviral drugs that block those parts, but not the parts involved in iron uptake.

The findings of Choe and colleagues also suggest that iron supplements may reduce the severity of New World virus infections. Past studies have shown that when the iron level in the body is low, the number of transferrin receptors in tissues increases. Consistent with these findings, Choe’s team found that New World arenaviruses infect cells more efficiently when iron levels are low, and that adding iron to cultured cells makes them less susceptible to infection. Choe notes that New World hemorrhagic fever outbreaks mostly occur in poor rural areas, where people are often deficient in micronutrients, including iron, possibly predisposing them to more severe infection when exposed to the rodents that carry the viruses.

Choe’s lab is now trying to find the cellular receptor for other viruses that cause hemorrhagic fever in humans. In 2003, Choe’s lab collaborated with Farzan’s lab to identify angiotensin converting enzyme2 (ACE2) as the receptor for the SARS virus.

The current study was funded by the National Institute of Allergy and Infectious Diseases.

For more information on viral hemorrhagic fevers, visit: http://www.cdc.gov/ncidod/dvrd/spb/mnpages/dispages/vhf.htm

For more information on arenaviruses, visit: http://www.cdc.gov/ncidod/dvrd/spb/mnpages/dispages/arena.htm

Children’s Hospital Boston is home to the world’s largest research enterprise based at a pediatric medical center, where its discoveries have benefited both children and adults since 1869. More than 500 scientists, including eight members of the National Academy of Sciences, 11 members of the Institute of Medicine and 10 members of the Howard Hughes Medical Institute comprise Children’s research community. Founded as a 20-bed hospital for children, Children’s Hospital Boston today is a 347-bed comprehensive center for pediatric and adolescent health care grounded in the values of excellence in patient care and sensitivity to the complex needs and diversity of children and families. Children’s also is the primary pediatric teaching affiliate of Harvard Medical School.

Children’s Hospital Boston
21 Autumn St., 2nd Fl.
Boston, MA 02115
United States
http://www.childrenshospital.org/

Strep, Psychiatric Symptoms Possibly Linked In Some Children

New research suggests that strep infections in children may increase involuntary movements and disruptive behaviors associated with some psychiatric disorders.

In an eight-month study of 693 children in a Florida public school system, University of Florida researchers found that shortly after the number of strep infections in the group increased, there was a corresponding rise in involuntary movements and disruptive behaviors symptoms that could indicate a neurological cause.

“During the fall months when there are more strep infections, after a short time lag, there are increased behavioral symptoms enough to indicate an association,” said Tanya Murphy, M.D., an associate professor of psychiatry in the College of Medicine. “We did not assess the children for particular neuropsychiatric disorders, so we’re not saying actual disorders were present in the children, but the symptoms were there.”

The research adds weight to the existence of PANDAS, short for Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcus. Some scientists think a host of problems such as tics, personality change, anxiety and obsessive-compulsive disorder may be triggered by strep infections in some children.

Scientists suspect group A streptococcal infections the kind that cause strep throat in some people but occur without symptoms in others may cause the body’s immune system to interact with brain cells that cause psychiatric symptoms in a small percentage of young patients.

In findings published this month in the Journal of Biological Psychiatry, UF researchers describe how they found an association between strep infections and neuropsychiatric symptoms within a group of students in a Florida school system. Previously, research in the PANDAS field focused on children already diagnosed with psychiatric disorders.

“We were looking for patterns of association in just a standard group of children who ranged in age from 3 to 12 years,” Murphy said. “We were seeing 693 kids once a month for eight months and made more than 5,000 observations.”

Throat cultures were collected to test for group A streptococcal infections while a clinician screened for tics and other involuntary movements of the fingers, wrists, arms, elbows and shoulders. In addition, as the children waited in line for their neurological screenings, the researcher made note of tic movements or any of nine categories of behaviors, ranging from fidgeting and hair-twirling to excessive touching and grimacing.

Analysis showed about 26 percent of children who had two or more strep infections displayed abnormal symptoms compared with 17 percent of children who were not infected or infected only once.

Strep throat and other group A strep infections are common in schools and environments where bacteria are easily spread. They are passed through direct contact with saliva or nasal discharge from an infected person, according to the National Institute of Allergy and Infectious Diseases.

Depending on the season strep peaks in December and January more than one in three children will be infected. Not all of the children will have sore throat symptoms, but they carry the bacteria and can infect others.

“The medical perspective has always been that the carrier states are fairly benign, but maybe they are not as benign as we thought,” Murphy said. “That’s not to suggest that these states are increasing children’s risk for rheumatic fever or other problems that can develop after an infection, but maybe there is a milder spectrum of effects that shouldn’t always be ignored.”

The research was funded in part through the National Institute of Mental Health. Other scientists involved include Sue Swedo, M.D., chief of the National Institute of Mental Health’s Pediatrics and Developmental Neuropsychiatry Branch, and Wayne Goodman, M.D., chairman of UF’s psychiatry department.

Determining whether strep truly triggers psychiatric disorders in some children will require further exploration. Scientists would next like to monitor the effect of strep treatment on psychiatric symptoms or observe whether a patient’s infection-fighting antibodies rise or fall in step with psychiatric symptoms.

“This is exactly the kind of study that was needed, a prospective evaluation to quantify the increased risk of neuropsychiatric or movement disorders following strep infections in the general pediatric population,” said Loren Mell, M.D., of the University of Chicago. Mell was part of a team that published findings in 2005 that showed strep infections were associated with increased risk of obsessive-compulsive disorder, Tourette’s syndrome or tic disorder in children who were already diagnosed with a psychiatric disorder.

“Further study to show prospectively that group A strep infections lead to neuropsychiatric disorders as determined by the Diagnostic and Statistical Manual of Mental Disorders criteria would help substantiate their findings,” Mell said. “Interestingly their results are similar to ours in the sense that having multiple infections appears to confer a much higher risk of these disorders.”

University of Florida Health Science Center
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AmeriHealth Expands Disease Management Program To Benefit Members With Crohn’s Disease

AmeriHealth today announced it has added the gastrointestinal disorder known as Crohn’s disease to its suite of disease management programs that serves more than 200,000 of its members.

AmeriHealth offers the broadest array of specialized, fully accredited disease management programs in the region. With the expansion of its disease management programs, the company now provides support and educational services to its members and their physicians for 22 chronic conditions.

Crohn’s disease is an incurable, chronic disorder that causes inflammation of the digestive or gastrointestinal (GI) tract. It most commonly affects the small intestine and colon, although it can involve any area of the GI tract. The cause of the disease is still unknown.

AmeriHealth’s Connections(SM) disease management program helps members with Crohn’s disease in several ways:

— Educates members about early symptom awareness, importance of complying with their therapy, and reducing risk factors.

— Reduces Crohn’s disease complications such as infections, fluid and electrolyte imbalance.

— Prevents certain complications from surgery and recovery period. (About three-fourths of Crohn’s disease patients have surgery during the course of the condition and almost half need repeat surgery).

“With Connections, members will find the tools and support to help them make informed health care decisions they feel good about, while strengthening their relationship with their doctors,” said Allan Goldstein, MD, vice president and regional medical director at AmeriHealth. “Crohn’s disease can be debilitating, but with information and specialized care, members can take control of their condition to take charge of their health and their lives.”

AmeriHealth’s suite of disease management programs provide support for 22 conditions – five common chronic diseases such as asthma and diabetes, offered through an alliance with Health Dialog; end-stage renal disease, in collaboration with RMS Disease Management Services; and 16 complex chronic diseases such as rheumatoid arthritis, multiple sclerosis, and the recently added Crohn’s disease, offered through Accordant Health Services, a Caremark company.

The Connections programs recently received national recognition from the Disease Management Association of America when they were awarded the “2006 Disease Management Leadership Award for Outstanding Health Plan” for innovative program design, successful plan implementation, and outstanding performance in measuring positive, proven health outcomes.

About AmeriHealth

AmeriHealth, Inc., a growing group of health care plans with its base in the states of New Jersey, Pennsylvania, and Delaware, has grown to more than 265,000 members since its inception in 1995.

AmeriHealth companies have been recognized for their commitment to providing high-quality products. AmeriHealth HMO of New Jersey and Delaware’s AmeriHealth HMO have received the highest possible accreditation by the national leader in HMO quality evaluation. The National Committee for Quality Assurance (NCQA) has awarded both HMOs “Excellent” status for meeting its rigorous evaluation standards.

About Accordant Health Services

Accordant Health Services, Inc., a Caremark company, provides health plans and employers with single-source health management solutions that lead to optimal health outcomes and greater satisfaction. A recognized leader in delivering disease management services for complex conditions, Accordant has leveraged the strengths of its disease management protocols with those of Caremark, which brings 10 years of common chronic disease management expertise to the marketplace.

Accordant empowers plan participants to make wise healthcare choices and adhere to their treatment regimens through a comprehensive approach, including disease management, HRA, wellness programs and case management. For more information about Accordant, visit http://www.accordant.net.

About Caremark Rx, Inc.

Caremark Rx, Inc. is a leading pharmaceutical services company, providing through its affiliates comprehensive drug benefit services to health plan sponsors and their plan participants throughout the U.S. The company’s clients include corporate health plans, managed care organizations, insurance companies, unions, government agencies and other funded benefit plans. In addition, Caremark is a national provider of drug benefits to eligible beneficiaries under the Medicare Part D program. The company operates a national retail pharmacy network with over 60,000 participating pharmacies, seven mail service pharmacies, the industry’s only FDA-regulated repackaging plant and 21 licensed specialty pharmacies for delivery of advanced medications to individuals with chronic or genetic diseases and disorders.

Additional information about Caremark is available at http://www.caremarkrx.com or in the company’s Forms 10-K, 10-Q and other SEC filings.

AmeriHealth, Inc.
http://www.accordant.net