ABERDEEN PROVING GROUND, Md. -- Scientists began noticing a drastic decline of bee colonies with no known cause in 2007. Many studies said that this decline of bees may put the country at risk of a food disaster. What few likely know is that a team of scientists at the U.S. Army Edgewood Chemical Biological Center utilized its own software to detect that a combination of both a virus and a fungi was the likely culprit.
Scientists with ECBC's Research & Technology Directorate's Point Detection Branch continue to enhance and expand this novel software suite of bioinformatics algorithms, which is capable of identifying biological microbes in various backgrounds without any prior knowledge of the sample. Utilizing data from a mass spectrometry-based proteomics system, the team can run any tandem mass spectral data through the software to provide statistical validation of its identity.
The innovative detection algorithm, known as Agents of Biological Origin Identifier, with two patents to date, is capable of providing automated identification of the sample contents from both pure cultures and mixture of microbes present in culture, environmental or biological matrices in far less time than traditional techniques--minutes versus days. When one puts this into the context of the recent letters laced with ricin that were sent to the president and other government officials, quick and accurate identification is imperative.
Mass spectrometry-based proteomics is a technique capable of providing the structured sequence information of a given protein. All proteins consist of amino acids arranged in a specific sequence; changing the arrangement generates a new protein. Mass spectrometry-based proteomics looks at all the digested protein fragments and peptides and determines the exact sequence arrangement of the peptides. It then correlates them to its original proteins that are present in a given sample, whether it's bacterial, viral or toxin.
"We often refer to this as a fishnet approach; with this technique, we collect everything and then sort through it to find what has biological meaning," said Rabih Jabbour, Ph.D., Research Chemist with the Point Detection Branch. "Unlike other techniques, we need no primers for a specific organism, just like with a fishnet you need no bait for a specific fish. We look at it all and then provide identification of microbes in a given sample."
The mass spectra signatures are then run through ABOid, which sorts out the information on a genomic level and translates it into proteomic output. Results are color-coded to highlight anything that could be considered a pathogen and whether it is toxic. For sequenced organisms, ABOid is capable of providing strain level identification, which is significant in that one can then reveal important diagnostic information for medical countermeasures.
ABOid utilizes a taxonomic classification approach to classify emerging or unknown--or unsequenced--microbes. If a specific bacteria or microbe is not sequenced, the team can use this approach to determine the closest near-neighbor to a given microbe. This complements polymerase chain reaction (PCR) technology and genomic sequencing, providing a critical first step to a genomics team to design an effective primer to sequence the bacteria to a specific target.
Algorithms are hardware independent, so they need only the data file from a mass spectrometer, not the sample itself, which eliminates the cost and logistics of shipping potentially toxic biological material. As cloud technology becomes more advanced, shipping costs will be reduced even further and the applications for ABOid will continue to expand.
"This significantly cuts down on the cost and red tape when shipping internationally or even across state lines, serving to further increase efficiency," said Mary Wade, Ph.D., Point Detection Branch Chief. "From a logistical standpoint, it's especially important when you consider soldiers or other personnel out in the field who can quickly send the data to a lab equipped with ABOid for quick analysis and accurate identification."
Another unique aspect of ABOid is that it can be tailored to meet specific needs. The Point Detection Branch narrowed the database to 104 viruses specific to the needs of the 20th Support Command, which is currently being trained on ABOid. The full database currently scans for more than 2,800 bacteria, 3,600 viruses and all known toxins, and more than 80 fungi and parasites--a significant growth since the development of ABOid.
Samir Deshpande, Ph.D., Research Bioinformatician with the Point Detection Branch, first began writing the code for ABOid in 2005. Through funding from the Defense Threat Reduction Agency, the Point Detection Branch was able to develop this software to become highly effective in identifying an unknown sample, and they were awarded the first-ever DTRA basic research award in 2008.
Since that time, the team continues to update the algorithms to ensure the software is compatible with updated mass spectrometry hardware as well as incorporating and curating newly sequenced organism continually.
"Since ABOid was created, there have been multiple advancements in mass spectrometry technology resulting in increased sensitivity and throughput," said Deshpande. "We are only bound by the sensitivity of the mass spec: the more sensitive the hardware, the more it can 'see,' which helps provide even better information for ABOid."
To date, ABOid has been has been utilized for multiple applications including detection of viruses, toxins and bacteria in various matrices, from buffers to more complex matrices such as blood. Through basic research funding from the U.S. Army In-House Laboratory Independent Research program, ABOid has been utilized to discriminate between pathogenic and non-pathogenic strains of E. coli, as well as evaluating the use of excreted proteins only for identification of bacteria.
ABOid also has applications in food safety, and the Point Detection Branch has collaborated with the U.S. Army Public Health Command for initial testing for salmonella in mashed potatoes and lettuce. PHC is focused on food safety for the Warfighter, looking to ensure rapid, accurate testing of any food source should a soldier become ill. The U.S. Army Public Health Command Region South, Food Analysis and Diagnostic Laboratory created the blinded food samples, some containing salmonella and some not, which were then shipped to ECBC for analysis. Because ABOid is a nonrestrictive detection technique, the system was able to accurately identify salmonella in each tainted sample, and also identified other background organisms present. That is a key factor when identifying any bacteria or toxins that may exist in a soldier's food.
With positive test results from the salmonella samples, ECBC and PHC are now looking to conduct testing for other potential toxins that could affect foodstuffs, such as ricin and botulinum toxin. There has been other interest in additional ABOid applications as well, such as the use of ABOid in outbreak scenarios, biosurveillance and bioforensic profiling.
ABOid will be among several systems tested as part of the new advanced technology demonstration - the Joint USFK (United States Forces Korea) Portal and Integrated Threat Recognition. The team will take its mass spectrometry equipment and the software to the Korean Peninsula next summer for field tests in conjunction with Contamination Avoidance, where it will be evaluated onsite for its ability to identify true and false positives. The team hopes that the opportunity will serve as a launching pad to eventually miniaturize, ruggedize and field the system, allowing the technology to be in the hands of one of the U.S. Army's most needed resources: the Warfighter.
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