Plants provide accurate low-cost alternative for diagnosis of West Nile Virus

October 24, 2012

While the United States has largely been spared the scourge of mosquito-borne diseases endemic to the developing world – including yellow fever, malaria and dengue fever – mosquito-related illnesses in the United States are on the rise. One pathogen of increasing national concern is an arbovirus known as West Nile. 

Now Qiang “Shawn” Chen, a researcher at Arizona State University’s Biodesign Institute and a professor in the College of Technology and Innovation, has developed a new method of testing for West Nile, using plants to produce biological reagents for detection and diagnosis.  Mosquito biting human Download Full Image

The new research, conducted by Chen and his colleagues at the Center for Infectious Diseases and Vaccinology, recently appeared in the Journal of Biomedicine and Biotechnology.

“One critical issue in WNV diagnosis concerns the difficulty of distinguishing WNV infection from other closely related diseases, such as St. Louis encephalitis and dengue fever, due to the cross-reactivity of antibodies among flaviviruses,” Chen says. “It is important to develop better diagnostic tools with enhanced accuracy for both treatment and diagnostic purposes.”   

Thus far, the 2012 outbreak of West Nile in the United States is on track to be one of the worst on record. According to the Center for Disease Control, 48 states have reported West Nile virus infections in people, birds or mosquitoes as of Oct. 9 of this year. 

To date, 4,249 cases of West Nile virus disease have been reported in humans, including 168 deaths. Of these cases 2,123 (50 percent) appeared in the more severe or neuroinvasive form of the disease, causing meningitis and encephalitis, while 2,126 cases were classified as non-neuroinvasive. 

These figures represent the highest number of West Nile cases reported to the CDC since 2003, with nearly 70 percent reported from eight states: Texas, California, Louisiana, Mississippi, South Dakota, Michigan, Oklahoma, and Illinois. Over a third of total cases have been reported from Texas.

The alarming upswing in West Nile cases coupled with their broad geographic distribution demand new techniques for both diagnosis and treatment. Chen and his colleagues have been exploiting the power of plant biotechnology to achieve these goals. 

Earlier, Chen’s group developed the first successful plant-derived therapeutic to combat West Nile post-infection, reporting their results in the Proceedings of the National Academy of Science. The current study advances efforts to create a diagnostic test for West Nile that will overcome barriers of existing methods, including limited accuracy, prohibitive cost and scalability. 

In nearly all cases, West Nile is transmitted to humans through the bite of an infected mosquito. Mosquitoes acquire the virus after feeding on infected birds. The virus then migrates to the mosquito's salivary glands, from which it may be injected into humans and animals. There, it can multiply and produce characteristic symptoms of West Nile disease. These may present as flu-like malaise, including fever and chills, headaches, fatigue and pain in muscles and joints. Symptoms typically last three to six days, but may persist for weeks.

In around 1 in 150 WNV cases, individuals develop infections of the brain (encephalitis) or surrounding tissue (meningitis), often producing severe headache, fever, stiffness, confusion, convulsions, coma, tremors, muscle weakness and paralysis. Those with neurologic involvement may require weeks of hospitalization and may suffer permanent health effects including muscle weakness and paralysis. Around 10 percent of people with WNV encephalitis die.

Faced with the growing threat of mosquito-borne epidemics, researchers like Chen stress the necessity of developing rapid, low-cost platforms for diagnosis of West Nile. Traditionally, cell cultures from serum, cerebrospinal fluid or tissues have been examined but the short viremic phase and low viral count of WNV in blood and spinal fluid limit the sensitivity and accuracy of such tests. Protein-based methods like ELISA have become standard tests for West Nile, yielding better results but at considerably higher cost and with limited scalability. 

In the current study, plants were exploited for their ability to produce large volumes of proteins that can be used for diagnostic testing. As Chen explains, proteins produced in this way traditionally require a lengthy time period before transgenic plant lines can be established. By contrast, the new method, which makes use of plant viral-based vectors like Tobacco Mozaic Virus and Gemini Virus, relies on the ability of plants to transiently express particular target genes, yielding the desired protein in one to two weeks.  

The technique provides the speed and flexibility of a bacterial gene expression system while permitting the posttranslational modifications of proteins afforded by mammalian cell culture approaches. 

Chen’s group used plant transient expression systems to produce two varieties of protein reagents useful for the detection and diagnosis of WNV – one a recombinant antigen and one a monoclonal antibody. High expression levels of both reagents were observed in two kinds of plants: Nicotiana benthamiana (a close relative of tobacco) and lettuce. The two reagents may be readily purified to greater than 95 percent and retain their native functionality and specificity.

The production of plant-derived antigens and monoclonal antibodies offers an attractive alternative to the use of mammalian, insect or bacterial cell cultures and demonstrates the capability of plants to provide accurate and flexible diagnostic reagents not only for WNV but a broad range of arboviruses affecting human health. 

“Our test will improve the accuracy of diagnosis, leading to the proper treatment of patients affected by WNV,” Chen says. “The plant-derived monoclonal antibody we examined is not only low-cost, but highly specific for WNV antigen and does not recognize antigens from other flaviviruses.” Chen further notes that application of this research will ultimately allow a broad range of WNV surveillance capabilities,  from clinical diagnosis to global distribution patterns in wild bird and mosquito populations. 

(Read the study)

West Nile Facts:

- The West Nile Virus (WNV) is an arbovirus belonging to the Flavivirus genus of the Flaviviridae family

- Mosquitoes that have acquired the virus from infected birds transmit WNV.

- First identified in the Eastern Hemisphere and widely distributed in Africa, Asia, Europe and the Middle East, WNV entered the Western Hemisphere in 1999, via New York City.

- Since its American debut, WNV has infected over 30,000 people with severe forms of the disease, with an untold number of additional cases remaining undiagnosed.

- Infection with WNV can be asymptomtic or can lead to West Nile fever or severe West Nile disease.

- Around 20 percent of people who become infected with WNV will develop West Nile fever, with symptoms including fever, headache, exhaustion and body aches, occasionally with a skin rash (on the trunk of the body) and swollen lymph glands. The illness can last from a few days to several weeks.

- The symptoms of severe WNV disease – also known as neuroinvasive disease, including West Nile encephalitis or meningitis or West Nile poliomyelitis – include headache, high fever, neck stiffness, stupor, disorientation, coma, tremors, convulsions, muscle weakness, and paralysis.

- Approximately 1 in 150 people infected with WNV develop a more severe form of disease. While serious illness can occur in people of any age, those over 50 as well as individuals with compromised immunity (including transplant patients) are at the highest risk for severe illness following WNV infection.

- The incubation time for WNV is typically 2 to 15 days. It is believed that infection confers lifelong immunity to the virus.

- This year (2012) has seen a serious uptick in West Nile cases and will go on record as among the most severe in terms of case numbers.

- According to Scientific American, a mild winter and a hot, dry summer have contributed to the rapid spread of WNV across the U.S. this year. Climate change has been implicated as a factor exacerbating the spread of WNV.

- The susceptibility to severe symptoms of WNV disease appears linked to genetic traits. Further, advanced age is one of the primary risk factors for severe neurological disease, long-term morbidity and death.

- Plant production for diagnostic protein reagents may easily be scaled up to suit existing need, as in the case of a sudden WNV epidemic.

- Plants are ideal protein bioreactors, due to their capacity to produce large volumes or proteins at low cost and their ability to make appropriate posttranslational modifications of proteins.

- Currently, there is no vaccine against WNV, nor effective therapeutics approved for human use. The best tactics of prevention are:

• Cover exposed skin with long pants and long-sleeved shirts, particularly during early morning or evening hours when mosquitoes are most active.

• Use insect repellent.

• Keep window screens in good repair.

• Try to reduce or eliminate areas of standing water where mosquitoes breed, including bird baths, wading pools, tire swings, etc.

Richard Harth

Science writer, Biodesign Institute at ASU


Study sheds light on Advanced Placement courses in Arizona schools

October 24, 2012

Advanced Placement (AP) courses offer students the opportunity to take rigorous, college-level classes in high school. Many colleges and universities offer college credit to students who earn a sufficiently high score on AP exams in one or more of 34 subjects. But access to the courses and student success on AP tests varies among Arizona’s urban and rural areas and racial/ethnic groups, according to a new study.

The study conducted by a faculty member in ASU’s Mary Lou Fulton Teachers College, assisted by a group of students pursuing their doctorate degrees in educational policy and evaluation, examines students’ access, participation, and success in AP courses in Arizona’s public high schools. Download Full Image

“We hope that our results help district leaders and policymakers better understand students’ participation in AP courses and test-taking as they consider the place AP courses have in a district’s curriculum and the allocation of resources,” said Jeanne M. Powers, an associate professor in Teachers College who was the lead author of “Advanced Placement Courses: Access, Participation, and Outcomes in Arizona,” recently published in Policy Points by ASU’s Morrison Institute of Public Policy.

Powers and her students analyzed 2009-2010 data from Arizona schools located in districts serving more than 3,000 students and a sample of smaller districts. Of those 172 public high schools, 80 percent offered at least one AP course. The majority of these schools are located in cities and suburbs. An average of 14 percent of students in these schools took at least one AP class. On average, Asian American and White students were overrepresented in AP courses, while American Indian, Black and Hispanic students were underrepresented.

The percentages of Arizona students taking and passing AP exams are below the national average. Of the Arizona students enrolled in AP courses, 54 percent took AP tests in at least one subject and 32 percent of enrolled students passed at least one AP test. While Hispanic students were the racial/ethnic group with the largest proportion of test takers, only 26 percent of the Hispanic students enrolled in AP courses passed at least one AP test. White and Asian students had lower rates of AP test-taking than Hispanic students, but passed AP tests at higher rates.

“During the process of putting the policy brief together, it was difficult to decide which findings to highlight,” said doctoral student Jesus Cisneros. “We suspected that disparities existed, but wanted to identify the communities most impacted by systematic inequalities.

“The argument could still be made that it is not just institutions with predominately minority student populations who have limited access to AP courses, but rather, institutions with predominately Hispanic student populations. Given the pervasive anti-immigrant sentiment encompassing our state, these findings are timely and significant. But we ultimately agreed that excluding other non-Hispanic minority students from our analysis would be a disservice to these populations, despite the potential impact of our findings,” Cisneros said.

“Unfortunately, I cannot say that I was shocked at the disparities we discovered in the reduced access for students who attend predominantly minority schools and rural schools, as this fits well within the larger body of literature regarding AP access,” said student Jessica Holloway-Libell. “I was, however, surprised to see the small percentages of students who take advantage of AP courses where courses are available. It would be interesting to explore the reasons behind this phenomenon.”

Powers and her student coauthors conclude the policy brief with a section addressing policy implications of their findings about AP courses and exams for Arizona’s schools and school districts.

“Having spent the past nine years in the field of education, I have seen firsthand the impact that education policies can have on the lives of students and teachers,” Holloway-Libell said. “I felt inclined to pursue a Ph.D. in this area to better understand the motivations behind, as well the intended and unintended consequences of such policies. I hope to one day contribute valuable research that helps policymakers develop policies that are in the best interest of students.”

Powers and the students worked together on researching their topic and writing up the analysis. Students wrote longer research papers related to the topic last fall as their finals; in the spring they continued working together to write the brief, which was submitted to the Morrison Institute in the fall.

“Having our doctoral students conduct research on local issues benefits the communities we serve,” said Suzanne Painter, director of the Division of Educational Leadership and Innovation in Teachers College. “I’m especially pleased that they did this work collaboratively, as they had to wrestle together with the complexities of research, and defend their methods and perspectives to each other.”

“The process of putting together a policy brief proved insightful for me as I learn to navigate the world of scholarship,” Holloway-Libell said. “One of the greatest challenges was the task of blending the voices, perspectives and personal values of seven individuals into one cohesive brief. After months of writing and rewriting, we arrived at what I consider a timely piece that adds important details to the overall discussion about AP courses in Arizona.”

The “Advanced Placement Courses: Access, Participation, and Outcomes in Arizona” study may be found online at