Download Report - Center for Food Safety Engineering - Purdue ...
Download Report - Center for Food Safety Engineering - Purdue ...
Download Report - Center for Food Safety Engineering - Purdue ...
You also want an ePaper? Increase the reach of your titles
YUMPU automatically turns print PDFs into web optimized ePapers that Google loves.
The <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
2005 - 2006 Research <strong>Report</strong><br />
“Collaborating to make our food safer”
The mission of the <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong><br />
<strong>Engineering</strong> is to develop new knowledge,<br />
technologies and systems <strong>for</strong> detection<br />
and prevention of chemical and microbial<br />
contamination of foods.<br />
Through CFSE, <strong>Purdue</strong> University positions<br />
itself as a national leader in multidisciplinary<br />
food safety research. Our multidisciplinary<br />
approach, including a strong<br />
engineering component, makes <strong>Purdue</strong><br />
University truly unique.
2005-2006 Research <strong>Report</strong><br />
2 Welcome from the Director<br />
• Message from Richard Linton, <strong>Center</strong> Director<br />
2 Message from USDA<br />
• Message from our Partnership with USDA-ARS<br />
3 Researchers developing food pathogen biosensors garner Agriculture Team Award<br />
3 <strong>Center</strong> Director captures IFT Myron Solberg Award<br />
4 <strong>Engineering</strong> of biosystems <strong>for</strong> the detection of Listeria Monocytogenes in foods<br />
• Michael R. Ladisch, Rashid Bashir, Arun Bhunia, J. Paul Robinson<br />
6 Multiplexed detection of pathogens using fl uorescence resonance energy transfer<br />
in spatial detection <strong>for</strong>mat<br />
• Bruce Applegate<br />
7 Optical <strong>for</strong>ward scattering <strong>for</strong> bacterial colony differentiation and identifi cation<br />
• Arun K. Bhunia, E. Dan Hirleman, J. Paul Robinson, Bartek Rajwa, Padmapriya Banda<br />
8 Multi-pathogen screening and/or confi rmation via microarray detection<br />
• Arun K. Bhunia, Mark Morgan, B.K. Hahm, Viswaprakash Nanduri, Shu-I Tu<br />
9 Multipathogen screening using immunomicroarray<br />
• Arun K. Bhunia, Viswaprakash Nanduri, Andrew Gehring<br />
10 Infrared sensors <strong>for</strong> rapid detection of select microbial foodborne contaminants<br />
• Lisa Mauer, Maribeth Cousin, Jay Gore, Jean Guard-Petter, Brad Reuhs, Sivakumar Santhanakrishnan<br />
11 Infrared sensors <strong>for</strong> rapid identifi cation of living vs. dead select microbial<br />
foodborne contaminants<br />
• Lisa Mauer, Maribeth Cousin, Jay Gore, Brad Reuhs<br />
12 Development of bioreporter-based chemical biosensor technology <strong>for</strong> the detection<br />
of chemical threat agents<br />
• David Nivens, Michael Franklin, Bruce Applegate, Carlos Corvalan<br />
13 Scientifi c publications and presentations<br />
16 <strong>Center</strong> staff<br />
Visit us @ www.cfse.purdue.edu
Welcome from the Director<br />
The <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong> (CFSE) at <strong>Purdue</strong> University, celebrates its 6th<br />
anniversary. It has been a very successful and exciting year <strong>for</strong> our interdisciplinary group.<br />
One of the key highlights was our biosensor team receiving the <strong>Purdue</strong> University College of<br />
Agriculture Team Award. This award is given each year to an interdisciplinary team that has<br />
demonstrated the most signifi cant research impact. Our partnership with USDA-ARS Eastern<br />
Regional Research Laboratory continues to breed success, leading to 19 peer-reviewed<br />
research publications, 21 presentations at national meetings, graduation of 5 Masters and Ph.D.<br />
students, and granting of 3 patents. This past summer, we were invited again to conduct a halfday<br />
program at the Annual Rapid Methods Workshop held at Kansas State University.<br />
Dr. Richard H. Linton<br />
Director of the <strong>Center</strong> <strong>for</strong><br />
<strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
Our research teams continue to work on emerging technologies to improve microbial and<br />
chemical detection. Detection systems are being developed <strong>for</strong> bacterial foodborne pathogens<br />
including Listeria monocytogenes, E. coli O157:H7, Campylobacter spp., Salmonella spp.,<br />
and a wide variety of chemical toxins. We are researching many detection technologies such<br />
as enzyme-linked immunosorbant assays, polymerase chain reactions, impedance-based<br />
microbiology, infrared spectroscopy, scanning microscopy, bioluminescense, and DNA/RNA<br />
probes. Some of the research breakthroughs include detection of live versus dead cells using<br />
infrared sensing devices, use of light scattering technology to produce images to distinguish<br />
foodborne pathogens from an agar plate, further development of a biochip <strong>for</strong> detection of<br />
Listeria monocytogenes, and signifi cant improvement of techniques to separate microorganisms<br />
from complex food systems.<br />
As our center grows, I continue to be amazed with the capabilities and talents of our scientists.<br />
To learn more about our center, please visit our web site at www.cfse.purdue.edu or feel free to<br />
contact me directly.<br />
Message from USDA<br />
Dr. Shu-I Tu<br />
Supervisory Research<br />
Chemist USDA-ARS,<br />
Eastern Regional<br />
Research <strong>Center</strong><br />
As the collaboration between the <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong> (CFSE) at <strong>Purdue</strong><br />
University and the Eastern Regional Research <strong>Center</strong> (ERRC) of the Agricultural Research<br />
Service (ARS) is entering its 7th year, I am grateful to witness the growth and maturation of<br />
this Congressional initiative. I am proud that the biosensor team of our <strong>Purdue</strong> colleagues was<br />
honored by winning the 2006 <strong>Purdue</strong> University College of Agriculture Team Award. Together,<br />
our <strong>Purdue</strong>-ERRC team has earned the reputation as a major contributor to the technology<br />
advancement of pathogen detection in food, as evident by the invitation from the International<br />
Workshop on Rapid Methods and Automation in Microbiology, to conduct a half-day symposium<br />
on molecular methodologies in August 2006. Furthermore, our team will have a dedicated issue<br />
of the Journal of Rapid Methods and Automation in Microbiology to report the progress of our<br />
collaboration team.<br />
In 2006, ERRC has continued to train visiting postdoctorates and graduate students from CFSE.<br />
Currently there are two visiting scientists from CFSE working at ERRC to develop phagedisplay<br />
antibodies, protein microarrays, and Salmonella detection methods using fi ber optics<br />
and time- resolved fl uorescence. We at ERRC valued the opportunity to be directly involved<br />
in and contribute to the development of future scientists in the food safety area. This year, the<br />
ERRC research team has started a 5-year research project to continue its ef<strong>for</strong>ts in biosensor<br />
research. This development assures the commitment by ARS to continue and strengthen the<br />
collaboration with CFSE <strong>for</strong> many more years.<br />
2<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Researchers developing food pathogen biosensors<br />
garner Agriculture Team Award<br />
An interdisciplinary team of scientists, who are inventing new ways to protect our food supply from potentially<br />
deadly food pathogens, has garnered the 2006 <strong>Purdue</strong> Agriculture Team Award.<br />
The Biosensor Detection Team’s<br />
research focuses on rapidly<br />
determining whether such microbes<br />
as Listeria monocytogenes or E.<br />
coli exist in food, particularly meat<br />
and milk products. The technologies<br />
the team has developed include an<br />
innovative biochip that analyzes<br />
very small amounts of food and<br />
does it faster and less expensively<br />
than current methods.<br />
The researchers also have found a<br />
way to take large samples of foods<br />
and concentrate the microorganisms into small volumes to inject onto the chip.<br />
“We have a number of different plat<strong>for</strong>ms in the team’s research because we have brought together<br />
many scientifi c disciplines,” said Arun Bhunia, a microbiologist in the Department of <strong>Food</strong> Science. “The<br />
interdisciplinary work not only has enabled us to produce these new technologies, but it also has helped us<br />
better teach and train undergraduate and graduate students in biology, microbiology and engineering. This<br />
award is a highlight and a result of the teamwork. We could not have achieved the same results if just one<br />
research specialty had been involved.”<br />
The biochip sensor, about the size of a fingernail, can determine if pathogen cells are present and whether<br />
they are dead or alive. Live cells cause disease, and it requires fewer than 10 cells to cause illness. The<br />
diagnostic method that includes the biosensor and the cell concentration technology has decreased detection<br />
time from two to three days to a few hours.<br />
“The technologies that the Biosensor Detection Team has developed are a major step in protecting food<br />
throughout the supply system from accidental or purposeful contamination by potentially deadly pathogens,”<br />
said Randy Woodson, Glenn W. Sample Dean of Agriculture. “This technology eventually will be available to<br />
many segments of the food-producing industry and will save time and money.”<br />
Approximately 76 millions cases of food-borne illness occur annually in the United States, according to the<br />
<strong>Center</strong>s <strong>for</strong> Disease Control and Prevention. This results in 325,000 people being hospitalized and 5,500<br />
deaths, and costs the health-care industry and individuals as much as $23 billion.<br />
Consumption of pathogen-contaminated food can cause meningitis, encephalitis (swelling of the brain), liver<br />
abscess, headache, fever and gastrointestinal problems. Listeria monocytogenes has a 20 percent fatality rate<br />
<strong>for</strong> those affected. The very young, people 60 and over, pregnant women and those with weakened immune<br />
systems are most at risk of being sickened by these pathogens.<br />
<strong>Center</strong> Director captures<br />
IFT Myron Solberg Award<br />
Dr. Richard Linton was the 2006 recipient of Myron Solberg Award presented at<br />
the Institute of <strong>Food</strong> Technologists (IFT) Annual Meeting in Orlando, Florida. The<br />
award honors an IFT member <strong>for</strong> providing world-class excellence in food science<br />
leadership in the establishment and successful development and continuation<br />
of industry, government, and academia cooperation. Linton was recognized <strong>for</strong><br />
his outstanding contributions <strong>for</strong> a) detection and control of foodborne pathogens,<br />
b) development of education and training programs <strong>for</strong> food manufacturing and<br />
retail food establishments, and c) exceptional service to science/industry boards,<br />
committees, and task <strong>for</strong>ces.<br />
3<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Ladisch, Bashir, Bhunia and Robinson<br />
<strong>Engineering</strong> of biosystems <strong>for</strong> the detection of Listeria Monocytogenes<br />
in foods<br />
Investigators: Michael R. Ladisch (Principal), Rashid Bashir, Arun Bhunia, J. Paul Robinson (College of <strong>Engineering</strong>) (College of Agriculture)<br />
Project Rationale<br />
Pathogenic bacteria cause 90% of reported foodborne illnesses.<br />
Listeria monocytogenes has emerged as one of the most<br />
important food pathogens, having a “zero tolerance” in ready-to<br />
eat processed (lunch) meats and dairy foods. This bacterium<br />
not only causes serious illness but also is lethal in infants,<br />
people over 60, and immune-compromised individuals.<br />
Current methods to detect this bacterium rely upon enrichment to<br />
increase the number of bacteria present in a sample. The food<br />
or food extract is incubated in special growth media <strong>for</strong> 12 to 24<br />
hours and the resulting culture is tested <strong>for</strong> L. monocytogenes<br />
using procedures that require an additional 3 to 24 hours.<br />
The food industry includes many small food processors and<br />
producers that do not have in-house microbiological laboratories<br />
<strong>for</strong> the purpose of testing <strong>for</strong> food pathogens. There<strong>for</strong>e, many<br />
companies send out samples <strong>for</strong> analysis. This adds up to<br />
another 24 hours to the time that elapses between when the<br />
food is sampled and the bacterium, if present, is detected. An<br />
overall time of 2 to 3 days typically elapses from when the food<br />
is sampled and the test results are available. The elapsed time,<br />
referred to as “time to result” or TTR, is problematic since some<br />
foods are consumed be<strong>for</strong>e test results would be available.<br />
Rapid and af<strong>for</strong>dable technologies to detect low numbers of L.<br />
monocytogenes cells directly from food, and which distinguish<br />
living from dead cells, are needed. This multi-disciplinary, multidepartmental<br />
research project is addressing the fundamental<br />
engineering and science required <strong>for</strong> development of<br />
microchip, bio-based assays that are transportable to the fi eld,<br />
useable in a manufacturing plant environment, and capable of<br />
rapidly detecting L. monocytogenes at the point of use. This<br />
research has the goals of (a) microscale detection of Listeria<br />
monocytogenes on a real-time or near real-time basis with a<br />
time-to-result of 4 hours, and, (b) reducing the time of culture<br />
steps with rapid cell concentration and recovery based on<br />
membrane technology.<br />
Our multidisciplinary research team is addressing the<br />
development, engineering and validation of such a microchip<br />
system that combines bioseparations technology <strong>for</strong><br />
rapid concentration with recovery of microbial cells and<br />
bionanotechnology to construct systems capable of interrogating<br />
fl uids <strong>for</strong> pathogens. Our approach is resulting in a technology<br />
plat<strong>for</strong>m capable of detecting other types of foodborne and<br />
medically relevant pathogens, even the focus of the research is<br />
on rapid detection of Listeria monocytogenes by a combination<br />
of technologies that will ultimately give a time to result of hours.<br />
Project Objectives<br />
Biochips are needed that are af<strong>for</strong>dable, capable of rapid<br />
detection of food pathogens, and easy to use by small food<br />
processors as well as major food companies. The goals and<br />
associated milestones of our research are:<br />
• Rapid concentration and recovery of microorganisms from<br />
food samples <strong>for</strong> subsequent interrogation of pathogens.<br />
• Sampling and conditioning fl uids containing the cells while<br />
maintaining their in<strong>for</strong>mation content (i.e., the molecules or<br />
cells that represent possible targets of the chip).<br />
• Transporting sample fl uids on and/or off the chip so that<br />
target microbes are captured and retained so that they can<br />
be probed <strong>for</strong> presence of pathogens.<br />
• Interfacing biological molecules (i.e., biomolecules) with<br />
electronic components.<br />
• Electronically detecting and amplifying biomolecular<br />
interactions between target and the biomolecules that <strong>for</strong>m<br />
the biorecognition components of the chip.<br />
• Achieving in vitro biospecifi city <strong>for</strong> the target molecule.<br />
• Interfacing biochip systems with electronic reading devices.<br />
4<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“Biochips are needed that are af<strong>for</strong>dable, capable of rapid detection of food pathogens, and<br />
easy to use by small food processors as well as major food companies.”
Ladisch, Bashir, Bhunia and Robinson<br />
Project Highlights<br />
The integration of cell concentration and recovery, sample<br />
introduction to the biochip, parallel detection of pH and<br />
conductivity, and improvements in specifi city and sensitivity<br />
have a common basis in rapid detection and quantitation<br />
of small differences in conductivity or changes in pH. This<br />
difference is maximized using a low conductivity buffer that will<br />
support cell viability and growth. The low conductivity growth<br />
medium (LCGM) that includes compounds such as tryptone,<br />
yeast extract, glucose, BSA and other constituents to yield a<br />
low conductivity of 1.2 mS. Proteins expressed by LCGM were<br />
identifi ed to include superoxide dismentase, thiol peroxidase,<br />
and unknown lipoproteins. These over-expressed proteins could<br />
be used as target proteins <strong>for</strong> development of new antibodies,<br />
and <strong>for</strong> direct or indirect measurement on-chip, thereby<br />
enhancing sensitivity. Overall, the validation of LCGM over the<br />
last year resulted in the practical impact of increasing sensitivity<br />
of on-chip detection of L. monocytogenes.<br />
indicate biological or chemical toxins. Such biomarkers may<br />
include proteins, peptides, low molecular weight metabolites,<br />
chemical or bio-toxins, and lipids, as well as nucleic acids. This<br />
is being studied in the context of on-chip pathogen detection and<br />
integration of preprocessing steps. The rapid concentration and<br />
recovery of the microorganisms has advanced with the use of<br />
hollow-fi ber membranes that are able to process extracts from<br />
foods that contain the microorganisms, and are amenable to use<br />
in a hands-off system that ultimately will lead to an automated<br />
instrument. A team approach is required by the complex<br />
interactions of the various components – both biological<br />
and electrical – of the detection system. These include<br />
dielectrophoresis and antibody-mediated selective capture of<br />
microorganisms in microfl uidic biochips. The multidisciplinary<br />
cooperation among the team has enabled signifi cant progress to<br />
be made in the integration of the various subsystems.<br />
Our interdisciplinary research approach has enabled us to learn<br />
about the biology of interactions between microorganisms and<br />
nutrient-rich foods; mechanisms of their transport and capture<br />
in microfl uidic systems; expression of biomarkers under<br />
environmental stress (i.e., conditions during food handling and/<br />
or sampling); and characteristics that impact their viability under<br />
environmental stress. The stress that the microorganisms may<br />
experience is related to changes in micro-environments during<br />
the course of sampling and rapid detection protocols.<br />
Initial runs with milk and vegetables have been carried out to<br />
begin examining rapid recovery of microorganisms internal<br />
to the biological tissue as found in various types of foods.<br />
Ultimately, the fundamental knowledge gained will apply to a<br />
broad range of samples (water, air, packages, animals, and<br />
people) <strong>for</strong> which fl uids are probed to rapidly assess the level<br />
of risk by interrogating these samples <strong>for</strong> biomarkers that<br />
“Listeria monocytogenes has emerged as one of the most important food pathogens,<br />
having a “zero tolerance” in ready-to eat processed (lunch) meats and dairy foods.”<br />
5<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Applegate<br />
Multiplexed detection of pathogens using fluorescence resonance energy<br />
transfer in spatial detection <strong>for</strong>mat<br />
Investigator: Bruce Applegate (Principal) (College of Agriculture)<br />
Project Rationale<br />
Inadvertent contamination of foods with harmful microorganisms<br />
can result in multiple problems including loss in productivity,<br />
expenses related to healthcare, investigation, litigation,<br />
destruction of vast quantities of agricultural products, and loss<br />
of human life. To streamline ef<strong>for</strong>ts in the circumvention of food<br />
poisoning-related incidents, FSIS requires a fully automated<br />
testing system <strong>for</strong> the rapid throughput analysis of foods <strong>for</strong><br />
contaminant pathogens (E. coli O157:H7, Salmonella, Listeria<br />
spp., etc.). Ideally, the testing plat<strong>for</strong>m will be a technicianoperated<br />
instrument that can simultaneously screen food<br />
samples <strong>for</strong> the presumptive presence of multiple bacteria<br />
and, if desired, confi rm their presence and characterize the<br />
pathogens through identifi cation of virulence-related or other<br />
genes. This approach would ultimately eliminate the need<br />
<strong>for</strong> the time-consuming conventional cell culture/isolation/<br />
confi rmation procedures currently used. The specific objective of<br />
this project is the development of a nucleic acid microarray <strong>for</strong><br />
the detection of multiple PCR products <strong>for</strong> the identifi cation of<br />
foodborne pathogens E. coli O157:H7, Salmonella spp. and L.<br />
monocytogenes, based on molecular beacon technology. This<br />
project leverages capabilities in DNA microarray technology<br />
to develop a gene-specifi c assay that does not require costly<br />
labeling and purifi cation methods to detect the presence of the<br />
target gene.<br />
Project Objectives<br />
• Construct an initial prototype gene array consisting of<br />
four marker genes <strong>for</strong> E. coli O157:H7 utilizing molecular<br />
beacon probes immobilized on a glass slide. (2005)<br />
• Determine hybridization conditions and evaluate<br />
the prototype beacon array utilizing amplifi ed target<br />
genes.(2005)<br />
• Contruct microarrays containing four targets from<br />
Salmonella spp. and L. monocytogenes from previously<br />
constructed target probe and amplicon sequences along<br />
with microarrays including all three sets of probes <strong>for</strong> the<br />
simultaneous detection of E. coli O157:H7, Salmonella spp.<br />
and L. monocytogenes. (2006)<br />
• Design and develop probe and amplicons <strong>for</strong><br />
Campylobacter, Clostridium, and S. aureus. (2006)<br />
• Develop and integrate a hybridization and PCR control into<br />
the array to provide quality assurance. (2006)<br />
• Construct arrays <strong>for</strong> multiple organisms and evaluate <strong>for</strong><br />
hybridization specifi city. (2006)<br />
• Evaluate previously developed multiplex PCR reactions<br />
<strong>for</strong> simultaneous amplifi cation of multiple pathogen targets<br />
utilizing the developed microarrays. (2006)<br />
Project Highlights<br />
This project focuses on the integration of the microarray<br />
work with the previously developed spatial array <strong>for</strong>mat. A<br />
DNA hybridization-based optical biosensor <strong>for</strong> the detection<br />
of foodborne pathogens was developed with virtually zero<br />
probability of a false negative signal. This portable, low-cost<br />
and real-time assaying biosensor utilizes the color-changing<br />
molecular beacon as a probe <strong>for</strong> the optical detection of the<br />
target sequence. The computer-controlled biosensor exploits the<br />
target hybridization-induced change of fl uorescence color due<br />
to the Förster (fl uorescence) resonance energy transfer (FRET)<br />
between a pair of spectrally shifted fl uorophores conjugated to<br />
the opposite ends of a beacon. Unlike the traditional fl uorophorequencher<br />
beacon design, the presence of two fl uorescence<br />
molecules allows one to actively visualize both hybridized and<br />
unhybridized states of the beacon. This eliminates false negative<br />
signal detection that is characteristic of the fl uorophore-quencher<br />
beacon <strong>for</strong> which bleaching of the fl uorophore or washout of<br />
a beacon is indistinguishable from the absence of the target<br />
DNA sequence. The two-color design allows us to quantify the<br />
concentration of the target DNA in a sample down to ≤0.5 ng/μl.<br />
The new design is suitable <strong>for</strong> simultaneous reliable detection of<br />
hundreds of DNA target sequences in one test run using a series<br />
of beacons immobilized on a single substrate in a spatial <strong>for</strong>mat.<br />
6<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“Inadvertent contamination of foods with harmful microorganisms can result in multiple<br />
problems including loss in productivity, expenses related to healthcare, investigation,<br />
litigation, destruction of vast quantities of agricultural products, and loss of human life.”
Bhunia, Hirleman, Robinson, Rajwa and Banda<br />
Optical <strong>for</strong>ward scattering <strong>for</strong> bacterial colony differentiation and<br />
identification<br />
Investigators: Arun K. Bhunia (Principal), E. Dan Hirleman, J. Paul Robinson, Bartek Rajwa, Padmapriya Banda (College of Agriculture, College of <strong>Engineering</strong>)<br />
Project Rationale<br />
Listeria monocytogenes and Escherichia coli are the major<br />
foodborne pathogens of concern in the United States. For the<br />
detection and evaluation of foods contaminated with Listeria<br />
monocytogenes or E. coli, USDA/FSIS recommends initial<br />
enrichment and subsequent plating on a selective agar media,<br />
which is often followed by further identifi cation procedures.<br />
These procedures are often time consuming and lengthy,<br />
taking more than 2-3 days. The present industrial demand is<br />
to increase the rapidity of the detection assays leading to<br />
strategies <strong>for</strong> decreasing bacterial contamination and thus<br />
reducing the economical loss. Our main objective was to<br />
reduce the time of identifi cation of these pathogens after<br />
plating, by developing a simple light scattering sensory method.<br />
The method has now been improved to differentiate among the<br />
different strains of Listeria, based on the varying patterns.<br />
There have been increasing foodborne illnesses, multiple<br />
outbreaks, product recalls, and loss of lives, resulting from<br />
pathogens in processed, ready-to-eat food products. Bacterial<br />
contamination in products not only puts the public at risk, it is<br />
costly to companies due to loss of production time, product<br />
recalls and liability.<br />
• To validate the technology by using naturally or<br />
deliberately contaminated food samples.<br />
• To analyze cellular composition, cell arrangement,<br />
refractive index and colony contents using electron<br />
microscopy, FT-IR or GC-MS.<br />
• To analyze the scatter signal images using ‘Standard<br />
feature extraction’ and ‘Moments of shape analysis’<br />
methods.<br />
Project Highlights<br />
The design and development of the BARDOT system was<br />
the major accomplishment during the year 2005-2006. This<br />
was a signifi cant accomplishment since it provided a plat<strong>for</strong>m<br />
<strong>for</strong> using a simple laser beam to scatter the bacterial colonies<br />
growing on an agar plate and simultaneously capturing and<br />
storing the images using a CCD camera attached to the<br />
instrument. The computer-stored images were used <strong>for</strong> image<br />
analysis. This system was able to differentiate genus Listeria,<br />
Salmonella and Vibrio with 89-98% accuracy. Furthermore,<br />
species within genus Listeria can be differentiated with 92-94%<br />
accuracy, Salmonella with 95-98% accuracy, and Vibrio with<br />
84-90% accuracy.<br />
Project Objectives<br />
• To improve the BARDOT (Bacteria Rapid Detection<br />
using Optical Scattering Technology) design, including<br />
supporting physics-based models, <strong>for</strong> more repeatability<br />
and maximum discrimination of <strong>for</strong>ward scattering<br />
signatures of colonies.<br />
• To acquire scatter images of colonies of select foodborne<br />
bacterial colonies including pathogens.<br />
• To analyze the bacterial colonies of different foodborne<br />
bacteria on non-selective and selective agar media.<br />
“the BARDOT system… provided a plat<strong>for</strong>m <strong>for</strong> using a simple laser beam to<br />
scatter the bacterial colonies growing on an agar plate and simultaneously<br />
capturing and storing the images…”<br />
7<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Bhunia, Morgan, Hahm, Nanduri and Tu<br />
Multi-pathogen screening and/or confirmation via microarray detection<br />
Investigators: Arun K. Bhunia (Principal), Mark Morgan, B.K. Hahm, Viswaprakash Nanduri, Shu-I Tu (College of Agriculture) (USDA-ARS, ERRC)<br />
Project Rationale<br />
<strong>Food</strong>borne disease is one of the most common causes of<br />
morbidity and mortality in the world, and more than 200<br />
known diseases are transmitted through food. In the United<br />
States there are about 76 million cases each year, of which<br />
325,000 require hospitalization and 5,000 die. The foodborne<br />
pathogens of concern are E. coli O157:H7, Salmonella, Listeria<br />
monocytogenes, Toxoplasma and Campylobacter. There<strong>for</strong>e,<br />
detection of these pathogenic bacteria during food processing<br />
and storage is crucial <strong>for</strong> the microbiological safety and<br />
prevention of possible outbreaks.<br />
Antibody-based detection methods are regarded as rapid and<br />
effi cient and are widely used in conventional ELISA and dipstick<br />
methods. In recent years, antibodies have been successfully<br />
used in biosensor tools <strong>for</strong> rapid detection. There<strong>for</strong>e,<br />
specifi city and avidity of a given antibody <strong>for</strong> the target bacteria<br />
is extremely important, specifi cally those originating from<br />
stressful environments of food. We have also learned that<br />
stress can affect antigen expression on bacterial cells thus<br />
affecting antibody-based detection.<br />
The goals of this project were to: (a) develop specifi c antibodies<br />
<strong>for</strong> Listeria monocytogenes, Salmonella enterica and E. coli<br />
O157:H7, (b) analyze effect of environmental and physiological<br />
stresses on antigen expression and antibody based detection,<br />
and (c) to develop antibody-based microarray system <strong>for</strong><br />
simultaneous detection of multi-pathogens.<br />
Project Objectives<br />
• Develop antibody specifi c <strong>for</strong> L. monocytogenes,<br />
Salmonella enterica and E. coli O157:H7.<br />
• Determine the effect of environmental and physiological<br />
stresses on antigen expression.<br />
• Develop sandwich ELISA <strong>for</strong> each pathogen.<br />
• Determine the selective enrichment media affecting the<br />
antibody-based detection of stress-exposed Listeria<br />
monocytogenes.<br />
• Development of pathogen enrichment and detection<br />
device (PEDD).<br />
Project Highlights<br />
Two polyclonal antibodies, PAb Lm404 and LmC369, were<br />
demonstrated to be specifi c <strong>for</strong> Internalin B (InlB) and<br />
actin polymerization protein (ActA) of L. monocytogenes,<br />
respectively. These antibodies could be potentially used <strong>for</strong><br />
specifi c detection of this bacterium. These antibodies showed<br />
differential expression of antigens in different enrichment<br />
broths: selective media (like BLEB, UVM and FB) suppressed<br />
PAb Lm 404 reactive InlB expression whereas only FB<br />
suppressed Lm C369 reactive ActA expression. PAb Lm404<br />
could be used only when bacteria are cultured in non selective<br />
media while PAb-Lm C369 could be used in an immunoassay<br />
with bacteria directly taken from selective enrichment broth.<br />
Surface localization of these two epitopes was confi rmed by<br />
immuno-electron microscopy.<br />
Once completed, this would allow development of a detection<br />
kit <strong>for</strong> multiple pathogens, thus saving time and money <strong>for</strong><br />
product testing and also helping regulatory agencies <strong>for</strong><br />
evaluation of a product <strong>for</strong> key food pathogens.<br />
8<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“<strong>Food</strong>borne disease is one of the most common<br />
causes of morbidity and mortality in the world…”
Bhunia, Hirleman, Robinson, Banda, Rajwa and Bayraktar<br />
Multipathogen screening using immunomicroarray<br />
Investigators: Arun K. Bhunia (Principal), Viswaprakash Nanduri, Andrew Gehring (College of Agriculture)<br />
Project Rationale<br />
Antibody-based methods are widely used <strong>for</strong> the detection<br />
of most pathogenic bacteria, and are regarded as rapid and<br />
effi cient. Their application to a conventional ELISA assay and<br />
further adaptation in modern biosensor tools shows promise in<br />
rapid detection. Most assays are developed <strong>for</strong> detection of a<br />
single target pathogen or toxin. There<strong>for</strong>e, it is very expensive<br />
to test <strong>for</strong> multiple pathogens from a single product because<br />
separate assay methods need to be used, thus adding cost<br />
and labor per test. Also, large laboratory space is required<br />
to per<strong>for</strong>m separate tests <strong>for</strong> each target pathogen because<br />
separate enrichment reagents and procedures need to be<br />
applied <strong>for</strong> different pathogens. If a single test is available that<br />
can detect multiple pathogens enriched in a single enrichment<br />
broth, this will not only reduce cost but also will be convenient<br />
and provide results in a short period of time. This would also<br />
benefi t the regulatory agencies when evaluating food products<br />
<strong>for</strong> key food pathogens.<br />
In the last decade, several rapid detection methods, such as<br />
antibody-based, nucleic acid-based, and biochemical-based,<br />
were developed. Even though these methods have shortened<br />
the analysis time compared to the conventional detection<br />
method, still we have to allow time <strong>for</strong> selective enrichment<br />
of samples prior to employing rapid detection methods. An<br />
antibody-based method such as ELISA requires at least 106<br />
CFU/ml <strong>for</strong> detection. Thus, to achieve that level of cells, it<br />
is important to use proper enrichment media <strong>for</strong> detection<br />
of foodborne pathogens. Furthermore, cell injury or stress<br />
encountered during food processing may affect cell numbers.<br />
Project Objectives<br />
The overall objective of this project is to develop<br />
immunomicroarray <strong>for</strong> concurrent detection of viable cells of<br />
three pathogens; L. monocytogenes, E. coli O157:H7 and<br />
Salmonella enterica.<br />
The specifi c objectives are:<br />
• Development of microarray assay in 96-well plate and<br />
glass slide using sandwich immunoassay <strong>for</strong> three<br />
pathogens.<br />
• Optimizing growth and enrichment of three pathogens<br />
(healthy or stressed) spiked in model food samples in a<br />
selective enrichment broth <strong>for</strong> use with microarray.<br />
Project Highlights<br />
Our group has developed a selective enrichment medium <strong>for</strong><br />
simultaneous growth and detection of Escherichia coli O157:<br />
H7, Listeria monocytogenes, and Salmonella. The media,<br />
designated SEL (Salmonella, E. coli, Listeria) was <strong>for</strong>mulated<br />
using Buffered Listeria Enrichment Broth (BLEB) base and<br />
various combinations of antibiotics: acrifl avine, cycloheximide,<br />
fosfomycin and nalidixic acid. Initial testing indicated that<br />
this medium supports the growth of E. coli O157:H7, L.<br />
monocytogenes and S. enteritidis well, and the growth rate of<br />
each is comparable to the respective selective enrichment broth<br />
such as modifi ed EC medium <strong>for</strong> E. coli, RV <strong>for</strong> Salmonella and<br />
FB <strong>for</strong> Listeria.<br />
Additionally, selective agents in media can delay growth and<br />
recovery of stressed or injured cells. Thus, currently used<br />
selective media may not be suitable <strong>for</strong> use with modern<br />
detection methods. Furthermore, current trends emphasize the<br />
detection of multiple targets with a single device. To achieve<br />
that, a medium that can allow selective enrichment of multiple<br />
pathogens is desirable.<br />
“If a single test is available that can detect multiple pathogens enriched<br />
in a single enrichment broth, this will not only reduce cost but also will be convenient<br />
and provide results in a short period of time.”<br />
9<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Mauer, Cousin, Gore, Guard-Petter, Reuhs and Santhanakrishnan<br />
Infrared sensors <strong>for</strong> rapid detection of select microbial foodborne<br />
contaminants<br />
Investigators: Lisa Mauer (Principal), Maribeth Cousin, Jay Gore, Jean Guard-Petter, Brad Reuhs, Sivakumar Santhanakrishnan<br />
(College of Agriculture, College of <strong>Engineering</strong>)<br />
Project Rationale<br />
Conventional detection methods take at least 24 to 48 hours to<br />
differentiate and identify microorganisms; there<strong>for</strong>e, measures<br />
taken to counteract food contamination must wait at least that<br />
long. To facilitate timely intervention measures, the food industry<br />
needs more rapid detection methods and a sensor able to<br />
accurately and rapidly identify low levels of microbial foodborne<br />
contaminants within food systems or culture media. We are<br />
investigating the effi cacy of infrared (IR) technology as a means<br />
<strong>for</strong> rapid detection of select bacterial pathogens. To accomplish<br />
this goal, we: (a) Created a library of Fourier-trans<strong>for</strong>m infrared<br />
(FT-IR) spectra of bacterial cell wall components and whole<br />
cells needed <strong>for</strong> pathogen identification and differentiation. (b)<br />
Developed FT-IR methods <strong>for</strong> identifi cation and quantifi cation<br />
of these pathogens from water, cultural media, and select<br />
foods. This included standardizing sampling procedures,<br />
quantifi cation methods, and spectral analysis procedures, as<br />
well as developing an overall chemometric approach <strong>for</strong> the<br />
analysis of FT-IR data. (c) Designed an IR sensor based on the<br />
most promising few-wavelength algorithms developed using<br />
FT-IR data generated from research activities in the fi rst two<br />
objectives.<br />
Project Objectives<br />
• Create a library of FT-IR spectra of bacterial cell<br />
wall components and whole cells (from Salmonella,<br />
Campylobacter jejuni, and Escherichia coli O157:H7)<br />
needed <strong>for</strong> cell identifi cation and differentiation.<br />
• Develop FT-IR methods <strong>for</strong> cell identifi cation and<br />
quantifi cation in water, cultural media, and foods.<br />
• Develop a limited wavelength approach <strong>for</strong> cell<br />
identifi cation.<br />
• Build and validate an IR sensor based on the most<br />
promising few-wavelength algorithm developed using FT-<br />
IR techniques selected in the fi rst two milestones.<br />
Project Highlights<br />
We successfully completed the development of sample<br />
preparation techniques <strong>for</strong> FTIR spectral collection and data<br />
analysis that are able to both quantify and identify Salmonella<br />
and E. coli O157:H7 from mixtures of bacteria in culture media<br />
and select food items. To develop this approach, we evaluated<br />
a variety of sample preparation methods, FTIR data collection<br />
methods, and analytical approaches <strong>for</strong> raw spectra. Further<br />
development of this approach enabled the design of a sensor<br />
that can be used in a production or retail facility to characterize<br />
a food sample as contaminated or free of select pathogenic<br />
bacteria in less time than current methods <strong>for</strong> detection.<br />
10<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“To facilitate timely intervention measures, the food industry<br />
needs more rapid detection methods…”
Mauer, Cousin, Gore and Reuhs<br />
Infrared sensors <strong>for</strong> rapid identification of living vs. dead select microbial<br />
foodborne contaminants<br />
Investigators: Lisa Mauer (Principal), Maribeth Cousin, Jay Gore, Brad Reuhs (College of Agriculture, College of <strong>Engineering</strong>)<br />
Project Rationale<br />
To keep the food supply safe, food production, processing,<br />
and retail establishments must be able to identify microbial<br />
foodborne pathogens, such as Salmonella, Campylobacter<br />
jejuni, and Escherichia coli O157:H7. The CDC estimates that<br />
annual foodborne-related outbreaks result in 76 million cases<br />
of illness, 325,000 hospitalizations, and 5,000 deaths.<br />
To facilitate timely intervention measures, the food industry<br />
needs rapid detection methods. Our group has developed<br />
such a system, using an infrared (IR) sensor, that is able<br />
to accurately identify low levels of microbial foodborne<br />
contaminants. In this phase of the study we have tested the<br />
fi eld instrument, and we have applied the technology to a<br />
second problem: distinguishing live bacterial cells from dead<br />
cells. To accomplish this goal, we developed FT-IR methods<br />
and analytical approaches to differentiate between living and<br />
dead cells of Salmonella spp. strains and E. coli O157:H7 in<br />
water, culture media, and select foods. This is important in both<br />
the determination of contamination potential in fresh products<br />
and the effi cacy of decontamination ef<strong>for</strong>ts. We also validated<br />
a miniature IR sensor <strong>for</strong> detecting the live bacteria. In this part<br />
of the project we addressed sample handling procedures, the<br />
time needed <strong>for</strong> detection, detection limits, and wavelength<br />
bands and algorithms appropriate <strong>for</strong> the sensor design.<br />
Project Objectives<br />
• Use an existing library of FT-IR spectra of bacterial cellwall<br />
components and whole cells of E. coli O157:H7 and<br />
E. coli K12, as well as Salmonella, <strong>for</strong> cell identifi cation<br />
and differentiation of live versus dead cells in water,<br />
cultural media, and foods. This milestone also involves<br />
the application and improvement of capture (fi ltration, etc),<br />
concentration, and enrichment techniques.<br />
• Apply a limited wavelength technique that has already<br />
been developed <strong>for</strong> cell identifi cation in the validation of<br />
a fi eld-size IR using the most promising few-wavelength<br />
algorithm developed during FT-IR techniques evaluations<br />
in the previous project.<br />
Project Highlights<br />
In the studies using variations on the selective and nonselective<br />
capture of live versus dead cells of E. coli O157:<br />
H7 and E. coli K12, and the subsequent spectral analyses,<br />
we showed that live cells could easily be distinguished from<br />
dead cells, especially when the cells were killed by the harsh<br />
techniques commonly used in the food industry (e.g., heat,<br />
salt, UV, etc). In contrast, the use of antibiotics that disrupt<br />
metabolism with out physical damage to the cell resulted in less<br />
distinct spectral separation of dead cells, but differentiation was<br />
still possible. Importantly, a relatively quick multi-step procedure<br />
allowed <strong>for</strong> a quantitative analysis of the relative abundance of<br />
dead versus live cells of E. coli O157:H7.<br />
“Our group has developed such a system, using an infrared (IR) sensor, that is able to<br />
accurately identify low levels of microbial foodborne contaminants.”<br />
11<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Nivens, Franklin, Applegate and Corvalan<br />
Development of bioreporter-based chemical biosensor technology <strong>for</strong> the<br />
detection of chemical threat agents<br />
Investigators: David Nivens (Principal), Michael Franklin, Bruce Applegate, Carlos Corvalan (College of Agriculture)<br />
Project Rationale<br />
Our nation must not only protect against environmental sources<br />
of pollution that can contaminate the food supply but also<br />
guard against deliberate acts of terrorism intended to degrade<br />
human health and/or weaken our economic base. The overall<br />
goal <strong>for</strong> our interdisciplinary research is the development of a<br />
core bioreporter-based chemical biosensor (BCB) plat<strong>for</strong>m<br />
<strong>for</strong> the eventual production of inexpensive biosensors to<br />
detect chemical agents that threaten our environment and<br />
our food supply. The BCB plat<strong>for</strong>m consists of an enclosure/<br />
microenvironment system that contains minimal nutrients,<br />
genetically-modifi ed bioreporters, and in some applications<br />
an analytical transducer. The technology exploits the abilities<br />
of living microorganisms: (i) to sense and react to chemical<br />
stimuli; (ii) to be genetically manipulated to contain reporter<br />
genes; and (iii) to <strong>for</strong>m biofi lms that promote survival.<br />
Enclosures/microenvironment systems will be developed to<br />
facilitate resuscitation of microorganisms from storage and<br />
sustain them within biofi lms in an optimal sensing state <strong>for</strong><br />
the detection of chemical agents. Our research involves the<br />
construction of bioreporters that are used in a novel biosensor<br />
confi guration to detect an organic pesticide (paraquat), a toxic<br />
metal (arsenic), and aromatic compounds (solvents). This<br />
research is expected to enable the development of BCB<br />
technology <strong>for</strong> rapid, selective and sensitive detection of many<br />
chemical threat agents. After core technology is developed,<br />
inexpensive application-specifi c devices will be designed <strong>for</strong><br />
potential use by farmers and/or scientists in a testing laboratory<br />
to identify environmental contamination or product tampering in<br />
both point-of-use and long-term monitoring applications.<br />
Project Objectives<br />
• Develop prototype enclosure(s) that contains a microenvironment<br />
that supports bio-reporting biofilms and in<br />
some applications contains a transducer(s) to facilitate<br />
rapid detection and long-term monitoring of biological<br />
responses to toxic compounds.<br />
• Develop biofi lm bioreporters <strong>for</strong> use in the enclosure/<br />
micro-environment <strong>for</strong> the detection of arsenic, paraquat,<br />
and aromatic solvents.<br />
• Combine constructed bioreporters with the prototype<br />
enclosure/micro-environments and obtain concentrationdependent<br />
bioreporter response data <strong>for</strong> detection of a<br />
chemical threat agent in food.<br />
• Use empirical data to develop models <strong>for</strong> understanding<br />
the nature of bacterial responses <strong>for</strong> improving the<br />
analytical per<strong>for</strong>mance of the biosensors.<br />
Project Highlights<br />
We have nearly completed the infrastructure <strong>for</strong> the core<br />
bioreporter-based sensor technology that will allow the<br />
development of application-specifi c sensors: (1) selection and<br />
testing of host strains, (2) constructing genetic systems that can<br />
be effi ciently and rapidly inserted into the genome of the host<br />
strain, (3) development of test systems to understand natural<br />
and confi ned biofi lms, (4) selection of appropriate transducers,<br />
and (5) development of mathematical models <strong>for</strong> testing.<br />
12<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“Our research involves the construction of bioreporters that are used in a<br />
novel biosensor confi guration to detect an organic pesticide (paraquat), a toxic<br />
metal (arsenic), and aromatic compounds (solvents).”
Scientific Publications and Presentations<br />
Peer Reviewed Journal Publications (2005-2006)<br />
• Bayraktar, B., Banada, P.P., Hirleman, E.D., Bhunia, A.K., Robinson, J.P., Rajwa. B. Bacterial phenotype identifi cation using Zernike<br />
moment invariants. Proceedings of the Society <strong>for</strong> PhotoOptical Instrumentation Engineers. v. 6080. p. 155-162 (2006).<br />
• Burgula, Y., Khali, D., Krishnan, S.S., Cousin, M.A., Gore, J.P., Reuhs, B. L., Mauer, L. J. Detection of E. coli O157:H7 and Salmonella<br />
Typhimurium Using Filtration followed by FT-IR Spectroscopy. Journal of <strong>Food</strong> Protection. 69:8. p. 1777-1784 (2006).<br />
• Chen, W-T., Hendrickson, R. L., Huang, C-P., Sherman, D., Geng, T., Bhunia, A. K., Ladisch, M. R. “Mechanistic Study of Membrane<br />
Concentration and Recovery of Listeria monocytogenes,” Biotechnol. Bioeng., 89, 263-273 (2005).<br />
• Chen, W-T., Geng, T., Bhunia, A. K., Ladisch, M. R. “Membrane <strong>for</strong> Selective Capture of the Microbial Pathogen Listeria<br />
monocytogenes,” AIChE J., 51(12), 3305-3308 (2005).<br />
• Hahm, B. K., Bhunia, A. K. Effect of environmental stresses on antibody-based detection of Escherichia coli O157:H7,<br />
Salmonella enterica subsp. Enteritidis and Listeria monocytogenes. Journal of Applied Microbiology. v. 100. p. 1017-1027<br />
(2006).<br />
• Huang, T. T., Taylor, D. G., Sedlak, M., Mosier, N. S., Ladisch, M. R. “Microfi ber-directed Boundary Flow in Press-fi t Microdevices<br />
Fabricated from Self-adhesive Hydrophobic Surfaces,” Analytical Chemistry, 77, 3671-3675 (2005).<br />
• Jedlica, S.S., McKenzie, J.L., Leavesley, S.J., Little, K.M., Webster, T.J., Robinson, J.P., Nivens, D.E., Rickus, J.L. Surface features of solgel<br />
derived silica infl uence protein con<strong>for</strong>mation and neuronal differentiation. Journal of Materials Chemistry. 16:3221-3230<br />
(2006).<br />
• Kim, H., Kane, M. , Kim, S., Dominguez, W., Applegate, B. Savikhin, S. A molecular beacon DNA microarray system <strong>for</strong> rapid<br />
detection of Escherichia coli O157:H7 that eliminates the risk of a false negative signal. Biosensors and Bioelectronics. In<br />
press (2006).<br />
• Kim, K.-P., Jagadeesan, B., Burkholder, K., Jaradat, Z.W., Wampler, J.L., Lathrop, A.A., Morgan, M.T., Bhunia, A.K. Adhesion<br />
characteristics of Listeria adhesion protein (LAP) – expressing Escherichia coli to Caco-2 cells and of recombinant LAP to<br />
eukaryotic receptor Hsp60 as examined in a surface plasmon resonance sensor. FEMS Microbiology Letters. v. 256. p. 324-<br />
332 (2006).<br />
• Kim, S. S., Huang, T. T., Fisher, T. S., Ladisch, M. R., “Effects of Carbon Nanotube Structure on Protein Adsorption,” IMECE 2005-<br />
2008 1395, Proceedings of IMECE 2005, 2005 ASME International Mechanical <strong>Engineering</strong> Congress and Exposition,<br />
Orlando, FL, November 5-100 (2005).<br />
• Kim, S., Kim, H., Reuhs, B.L., Mauer, L.J. Differentiation of outer membrane proteins from Salmonella enterica serotypes using<br />
Fourier trans<strong>for</strong>m infrared spectroscopy and chemometrics. Letters in Applied Microbiology. v. 42. p. 229-234 (2006).<br />
• Kim, S., Burgula, Y., Ojanen-Reuhs, T., Cousin, M.A., Reuhs, B.L., Mauer, L.J. Differentiation of crude lipopolysaccharides from<br />
Escherichia coli strains using Fourier trans<strong>for</strong>m infrared spectroscopy and chemometrics. Journal of <strong>Food</strong> Science. v. 71. p.<br />
57-61 (2006).<br />
• Lim, K. S., Chang, W.J., Koo, Y.M., Bashir, R., “Reliable Fabrication Method of Transferable Micron Scale Metal Pattern <strong>for</strong><br />
Poly(dimethylsiloxane) Metallization”, Lab. Chip. v. 1. 578 – 580 (2006).<br />
• Yang, L., Banada, P.P., Liu, Y.S., Bhunia, A.K., Bashir, R. Conductivity and pH dual detection of growth profi le of healthy and<br />
stressed Listeria monocytogenes. Biotechnology and Bioengineering. v. 92. p. 685-693 (2005).<br />
• Yang, L., Banada, P., Chatni, M. R., Lim, K, Ladisch, M., Bhunia, A., Bashir, R. “A MultiFunctional Micro-Fluidic System <strong>for</strong><br />
Dielectrophoretic Concentration Coupled with Immuno-Capture of Low Number of Listeria monocytogenes”, Lab on a chip,<br />
appeared online. DOI: 10.1039/b607061m (2006).<br />
“Our partnership with USDA-ARS Eastern Regional Research Laboratory continues to<br />
breed success, leading to 19 peer-reviewed research publications, 21 presentations at national<br />
meetings, graduation of 5 Masters and Ph.D. students, and granting of 3 patents.”<br />
13<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Scientific Publications and Presentations<br />
Abstracts <strong>for</strong> Major Papers/Posters Presented (2005-2006)<br />
• Bashir, B. “BioMEMS and Bionanotechnology and Applications to Diagnostics”, Plenary Session Monday Oct 31st, Topical<br />
Conference “Biomedical Applications of Nanotechnology (Bionanotechnology)”, AIChE Annual Meeting. Cincinnati, OH. 2005<br />
(invited).<br />
• Bashir, R. BioMEMS and Bionanotechnology and Applications to Diagnostics, in N.A. Peppas and J.Z. Hilt, eds., “Advances in<br />
Bionanotechnology”, pp. 1-5, AIChE, New York, NY. 2005 (invited).<br />
• Bayraktar, B., Banada, P. P., Hirleman, E. D., Bhunia, A. K., Robinson, J. P., Rajwa, B. Feature Extraction, Identifi cation and<br />
Classifi cation of Listeria colonies from light scatter patterns. 14th Annual GLIIFCA Meeting. 2005.<br />
• Bayraktar, B., Banada, P.P., Hirleman, E D, Bhunia, A.K., Robinson, J.P., Rajwa, B. Feature extraction from light-scatter patterns of<br />
Listeria colonies <strong>for</strong> identifi cation and classification. Journal of Biomedical Optics. 2006. v. 11 paper no. 034006.<br />
• Burgula, Y., Cousin, M.A., Reuhs, B.L., Mauer, L.J. Differentiation of Live vs. Dead cells of E. coli O157:H7 based upon incubation<br />
and immunomagnetic separation. Institute of <strong>Food</strong> Technologists’ Annual Meeting and <strong>Food</strong> Expo. Orlando, FL. 2006.<br />
• Burgula, Y., Cousin, M.A., Applegate, B., Linton, R., Reuhs, B.L., Mauer, L.J. Effects of processing treatments on FT-IR based<br />
classifi cation of dead E. coli K12 cells in comparison to live cells. Institute of <strong>Food</strong> Technologists’ Annual Meeting and <strong>Food</strong><br />
Expo. Orlando, FL. 2006.<br />
• Burkholder, K.M., Bhunia, A.K. Interaction of Salmonella with cultured intestinal cell lines during heat stress. American Society<br />
<strong>for</strong> Microbiology General Meeting. 2005. p. 445. Abstract No. P-041.<br />
• Gomez, R., Bashir, R. “Microscale Impedance-Based Detection of Bacterial Metabolism”, in Encyclopedia of Rapid<br />
Microbiological Methods, Volume 3, Series Editor, Michael J. Miller, Davis Healthcare International Publishing (DHI). 2006.<br />
pp. 333-362.<br />
• Huff, K., Banada, P.P., Bayraktar, B, Bae, E, Rajwa, B, Robinson, J.P., Hirleman, E.D. Bhunia, A.K. Detection and Identifi cation of<br />
<strong>Food</strong>borne Pathogens in Genus and Species Levels Using a Non-invasive Modifi ed Light Scatterometer-BARDOT. American<br />
Society <strong>for</strong> Microbiology Annual Meeting. 2006. Abstract no. P-075.<br />
• Jagadeesan, B., Burkholder, K., Wampler, J.L., Bhunia, A.K. Interaction of Listeria adhesion protein (LAP) with human Hsp60 on<br />
the surface of stressed epithelial cells. American Society <strong>for</strong> Microbiology General Meeting. 2006. p. 48. Abstract No. B-103.<br />
• Kwan S.L., Chang, W.J., Koo, Y.M., Bashir, R. “Embedding Microscale Metal Patterns In Polydimethylsiloxane Substrate” Ninth<br />
International Conference on Miniaturized Systems <strong>for</strong> Chemistry and Life Sciences (μTAS), Boston Marriott Copley Place,<br />
Boston, Massachusetts, USA October 9th - 13th, 2005.<br />
• Lathrop, A.A., Bhunia, A.K. Expression of Listeria monocytogenes InlB and ActA surface proteins under different growth media.<br />
American Society <strong>for</strong> Microbiology General Meeting. 2005. p. 446, Abstract No. P-046.<br />
• Paarlberg, K., Burgula, Y., Cousin, M.A., Reuhs, B.L., Mauer, L.J. Development of a FT-IR spectral library <strong>for</strong> select microorganisms.<br />
Institute of <strong>Food</strong> Technologists’ Annual Meeting and <strong>Food</strong> Expo. Orlando, FL. 2006. Abstract.<br />
• Shen, X., Nivens, D.E., Corvalan, C.M. Predicting analytical response in bioreporter-based sensors <strong>for</strong> food and agriculture<br />
systems. Institute <strong>for</strong> <strong>Food</strong> Technologists, Orlando, FL. 2006. Session number 020B.<br />
14<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
“At the <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>, we direct our
Thesis/Dissertations (2005-2006)<br />
• Burgula, Y., “Detection of select foodborne pathogens using FT-IR spectroscopy,” Ph.D. Dissertation. 2006. <strong>Purdue</strong><br />
University. 294p.<br />
• Chen, W., Biomedical <strong>Engineering</strong>, “<strong>Engineering</strong> and Analysis of Immobilized Enzyme System in Microfl uidic Device,” Ph.D.<br />
Dissertation. 2006. <strong>Purdue</strong> University. 135 p.<br />
• Davis, K.D. “Assessment of molecular virulence gene profi ling and antibodies <strong>for</strong> rapid detection of pathogenic Escherichia<br />
coli isolates.” Ph.D. Dissertation. 2005. <strong>Purdue</strong> University. 138 p.<br />
• Lathrop, A.A. “Development of Listeria monocytogenes specifi c antibodies using a proteomic/genomics approach and<br />
expression of antibody-specifi c antigens InlB and ActA under different environments.” Ph.D. Dissertation. 2005. <strong>Purdue</strong><br />
University. 142 p.<br />
• Nagel, A.C. “Development and analysis of bioreporters in biofi lms,” Masters Thesis. 2006. <strong>Purdue</strong> University. (In Progress)<br />
Books and Book Chapters (2005-2006)<br />
• Amass, S.F., Bhunia, A.K., Chaturvedi, A.R., Dolk, D.R., Peeta, S., Atallah, M.J. editors. <strong>Purdue</strong> University Press, West Lafayette, IN.<br />
Advances in Homeland Security Vol. 1. The Science of Homeland Security, 2006. p. 109-149.<br />
• Bhunia, A.K. Detection of signifi cant bacterial pathogens and toxins of interest in homeland security.<br />
Invited Lectures and Seminars (2005-2006)<br />
• Bhunia, A.K. Bacterial Pathogen Detection: Micro/Nano-Technology Approaches. Indian Institute of Chemical Biology,<br />
Calcutta, West Bengal, India (Dec 21, 2005).<br />
• Bhunia, A.K. Listeria monocytogenes Pathogenesis: Intestinal Phase of Infection. Indian Institute of Technology, Guwahati,<br />
Assam, India (Dec 5, 2005).<br />
• Bhunia, A.K. Microbiology Meets Nanotechnology. Indian Institute of Technology, Guwahati, Assam, India (Dec 5, 2005).<br />
• Bhunia, A.K. Optical immunosensors and cell-based detection <strong>for</strong> Listeria monocytogenes. International Rapid Methods<br />
Workshop, Kansas State University, Manhattan, KS (July, 2006).<br />
• Bhunia, A.K. Pathogenesis of Listeria monocytogenes and its detection strategies using micro/nano sensors. Department of<br />
Biology, Ball State University, Muncie, Indiana (Nov 4, 2005).<br />
• Ladisch, M. R., “Microfl uidic Characterization of Press Fit Microdevices,” University of Akron, Akron, OH (October 21, 2005).<br />
• Ladisch, M. R., “Rapid Prototyping of Microfl uidic Separation Plat<strong>for</strong>ms,” North Carolina State University (February 3, 2006).<br />
Patents Granted (2005-2006)<br />
• Bhunia, A.K., Hahm, B.K., Morgan, M.T. Pathogen enrichment detection device and methods of use. Ref. No. 64303.P1US<br />
(2005).<br />
• Hirleman, E.D., Guo, S., Bhunia, A.K., Bae, E. System and method <strong>for</strong> rapid detection and characterization of bacterial colonies<br />
using <strong>for</strong>ward light scattering. Ref. No. 64142.00.US (2005).<br />
• Ladisch et al, Cell Concentration and Pathogen Recovery, 11/081 378 (2005).<br />
ef<strong>for</strong>ts toward detecting problems and protecting consumers.”<br />
15<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
<strong>Center</strong> Staff<br />
Dr. Richard H. Linton<br />
Director<br />
765.494.6481<br />
linton@purdue.edu<br />
Staff<br />
Kevin T. Hamstra, Multimedia Technical Specialist<br />
• 765.496.3833 • khamstra@purdue.edu<br />
Kiya A. Smith, <strong>Center</strong> Coordinator<br />
• 765.496.3827 • kiya@purdue.edu<br />
Dr. W. R. (Randy) Woodson, Dean of Agriculture<br />
• 765.494.8391 • woodson@purdue.edu<br />
Dr. Shu-I Tu<br />
Supervisory Research Chemist<br />
215.233.6466<br />
stu@arserrc.gov<br />
USDA-ARS<br />
Dr. Jeffrey Brewster • 215.233.6447 • jeffrey.brewster@ars.usda.gov<br />
Dr. John P. Cherry • 215.233.6595 • john.cherry@ars.usda.gov<br />
Dr. Pina Fratamico • 215.233.6525 • pina.fratamico@ars.usda.gov<br />
Dr. Andrew Gehring • 215.233.6491 • andrew.gehring@ars.usda.gov<br />
Dr. Peter Irwin • 215.233.6420 • peter.irwin@ars.usda.gov<br />
Dr. James A. Lindsay • 301.504.4674 • jal@ars.usda.gov<br />
Dr. John B. Luchansky • 215.233.6620 • john.luchansky@ars.usda.gov<br />
Dr. George Paoli • 215.233.6671 • george.paoli@ars.usda.gov<br />
Dr. Gary Richards • 302.857.6419 • grichards@errc.ars.usda.gov<br />
Dr. Christopher Sommers • 215.836.3754 • chris.sommers@ars.usda.gov<br />
Dr. Howard Zhang • 215.233.6583 • howard.zhang@ars.usda.gov<br />
Dr. Bruce Applegate<br />
PI<br />
765.496.7920<br />
applegab@purdue.edu<br />
Co-PIs<br />
Dr. Michael Kane • mdkane@purdue.edu<br />
Dr. Lynda Perry • lperry@purdue.edu<br />
Staff / Graduate Students<br />
Preciaus Heard • pheard@purdue.edu<br />
Dr. Arun K. Bhunia<br />
PI<br />
765.494.5443<br />
bhunia@purdue.edu<br />
Co-PIs<br />
Padmapriya Banada • 765.496.3826 • pbanada@purdue.edu<br />
Bulent Bayraktar • 765.494.0757 • bayrakta@ecn.purdue.edu<br />
Andrew Gehring • agehring@errc.ars.usda.gov<br />
B. K. Hahm • 765.496.7356 • hahm@purdue.edu<br />
E. Dan Hirleman • 765.494.5688 • e.daniel.hirleman.1@purdue.edu<br />
Mark Morgan • 765.494.1180 • mmorgan@purdue.edu<br />
Viswaprakash Nanduri • vnanduri@purdue.edu<br />
Bartek Rajwa • 765.494.0757 • rajwa@fl owcyt.cyto.purdue.edu<br />
J. Paul Robinson • 765.494.6449 • jpr@fl owcyt.cyto.purdue.edu<br />
Shu-I Tu • 215.233.6466 • stu@errc.ars.usda.gov<br />
Staff / Graduate Students<br />
EuiWon Bae • 765.494.4762 • baee@ecn.purdue.edu<br />
Pratik Banerjee • 765.496.7356 • pbanerje@purdue.edu<br />
Karleigh Huff • khuff@purdue.edu<br />
Hyochin Kim • 765.496.7354 • kim399@purdue.edu<br />
16<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
Dr. Michael Ladisch<br />
PI<br />
765.494.7022<br />
ladisch@purdue.edu<br />
Co-PIs<br />
Padmapriya Banada • 765.496.3826 • pbanada@purdue.edu<br />
Rashid Bashir • basher@purdue.edu<br />
Arun Bhunia • 765.494.5443 • bhunia@purdue.edu<br />
Xingya Liu • 765.494.7052 • xingya@purdue.edu<br />
Nathan Mosier • 765.494.7025 • mosiern@purdue.edu<br />
J. Paul Robinson • 765.494.6449 • jpr@fl owcyt.cyto.purdue.edu<br />
Miroslav Sedlak • 765.494.3699 • sedlak@purdue.edu<br />
Staff / Graduate Students<br />
Ben Baker • babaker@purdue.edu<br />
Adam Burton • amburton@purdue.edu<br />
Jeremiah Bwatwa • jbwatwa@purdue.edu<br />
Katherine Gregory • klgregor@purdue.edu<br />
Balamurugan Jegadeeshan • baa@purdue.edu<br />
Mark Kim • mjkim@purdue.edu<br />
Yi-Shao Liu • yishao@purdue.edu<br />
Brandon Manier • bmanier@purdue.edu<br />
Chris (Heyjin) Park • 765.494.7031 • parkhc@purdue.edu<br />
Alex Serafin • ajserafi @purdu.edu<br />
Hunter Vibbert • hvibbert@ecn.purdue.edu<br />
Angela Valadez • 765.496.3824 • valadeza@purdue.edu<br />
Dr. Lisa Mauer<br />
PI<br />
765.494.9111<br />
mauer@purdue.edu<br />
Staff / Graduate Students<br />
Yashodar Burgula • 765.496.3805 • yash@purdue.edu<br />
Dr. David E. Nivens<br />
PI<br />
765.494.0460<br />
dnivens@purdue.edu<br />
Co-PIs<br />
Bruce Applegate • 765.496.7920 • applegab@purdue.edu<br />
Carlos Corvalan • 765.494.8262 • corvalac@purdue.edu<br />
Michael Franklin • 406.994.5658 • umbfm@montana.edu<br />
Staff / Graduate Students<br />
Bonnie Co • 765.496.1805 • cob@purdue.edu<br />
Claudia Ionita • 765.496.7354 • cionita@purdue.edu<br />
Aaron Nagel • 765.496.3832 • acnagel@purdue.edu<br />
David Schroeder • 765.496.1805 • dlschroe@purdue.edu<br />
Xinyuu Shen • 765.496.3825 • shenx@purdue.edu<br />
17<br />
<strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>
It is the policy of the <strong>Purdue</strong> University <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong>, that all persons shall have equal<br />
opportunity and access to the programs and facilities without regard to race, color, sex, religion, national origin,<br />
age, marital status, parental status, sexual orientation or disability.<br />
<strong>Purdue</strong> University is an Affirmative Action employer.<br />
The <strong>Center</strong> <strong>for</strong> <strong>Food</strong> <strong>Safety</strong> <strong>Engineering</strong><br />
<strong>Purdue</strong> University<br />
<strong>Food</strong> Science Building<br />
745 Agriculture Mall Drive<br />
West Lafayette, IN 47909<br />
Non-profit<br />
Organization<br />
U.S. Postage<br />
PAID<br />
<strong>Purdue</strong><br />
University