New detection technology identifies bacteria, viruses, other organisms within 24 hours


The pig virus, porcine circovirus-1 (PCV-1), was unexpectedly found in GlaxoSmithKline’s Rotarix vaccine, which is used to prevent diarrhea in babies. “One result of this research is that it demonstrates how modern technologies could change and drastically improve product safety,” Slezak said.

While product safety rules require demonstrating that a list of known contaminants is not present, Slezak said the use of modern advances in DNA sequencing and arrays would allow manufacturers to identify the potential presence of contaminating biological material present in quantities large enough to be of potential concern. “For each bacteria or virus that has been sequenced anywhere in the world, we have several dozen squares on the checkerboard that will identify sequences from that organism,” Slezak explained.

Currently, Slezak’s team is testing a next-generation LLMDA that boasts 2.1 million probes. This version contains probes representing about 178,000 viral sequences from some 5,700 viruses, and about 785,000 bacterial sequences from thousands of bacteria.

The latest LLMDA version also encompasses fungi and protozoa – with probes representing about 237,000 fungal sequences from thousands of fungi and about 202,000 protozoa sequences from 75 protozoa.

As a screening tool, Slezak sees the LLMDA as occupying niche roles between PCR machines and sequencing. The LLMDA process starts with the purification of DNA or RNA from a sample, such as sputum or blood. The sample is next labeled with a fluorescent dye and hybridized on the microarray at 42 degrees C or about 107.6 degrees Fahrenheit. In turn, a fluorescent scanner and analysis software are used to detect the probes that have lit up, identifying the presence of viral or bacterial sequences.

The Livermore team plans to update probes on the array with new sequences of bacteria, viruses and other microorganisms from GenBank and other public databases about once per year, in addition to using sequences obtained from collaborators for their probes.

LLNL’s current collaborators include the University of California, San Francisco; the Blood Systems Research Institute; the University of Texas Medical Branch (Galveston); the National Institute for Public Health and the Environment of Bilthoven, the Netherlands; the Statens Serum Institut of Copenhagen, Denmark; the University of California, Davis; Imigene; the U.S. Food & Drug Administration; and the Marine Mammal Center of Sausalito, California.

A computer scientist and the team’s leader, Slezak came up with the idea for the LLMDA in 2003. His team includes biologist Crystal Jaing, who leads the microarray lab work and manages the collaborations; bioinformaticist Shea Gardner, who designed the array; biostatistician Kevin McLoughlin, who designed the analysis software; and James B. Thissen, who performs the microarray experiments.