Gene Sequencing Added to Microbial Identification Options
The use of genotypic tests to identify the source of microbial contamination is changing the pharmaceutical microbiology laboratory in much the same way that DNA testing has changed the forensic laboratory. Globally, many regulatory agencies now consider DNA-based techniques to be more accurate than phenotypic methods. Lancaster Laboratories is expanding identification options by offering gene sequencing through its European facility. In addition, the recent installation of software upgrades for ribotyping and fatty acid analysis (FAME) augments their existing identification capabilities.
Lancaster Laboratories Europe, located in Dungarvan, Ireland, is currently using the MicroSEQ® Microbial Identification System manufactured by Applied Biosystems to perform gene sequencing. For this analysis, DNA is extracted from the cell culture, amplified by Polymerase Chain Reaction (PCR) and then sequenced. The resulting genome is compared to a database to identify the DNA. The bacterial library contains over 1800 species, and the system is also capable of analyzing yeasts and molds by comparison with a library of fungal species. This automated technique is more accurate and reproducible than traditional manual methods, which are highly dependent on analyst skill and consistency. In order to offer this testing to US based clients, Lancaster Laboratories will use a validated process for preparing cultures and shipping the resulting DNA to their Europe location for gene sequencing.
Lancaster Laboratories also continues to perform characterization of microorganisms to the strain level using their DuPont Qualicon RiboPrinter®. This genotypic analysis is a powerful analytical tool that creates a genetic snapshot of the organism, which is then compared to the recently expanded database of RiboPrint patterns for accurate identification. Because each organism yields a very specific pattern, the Riboprinter® is often the instrument of choice for investigation of positive results for sterility and microbial limits testing. Even patterns that are not identified as a match in the database can be useful to track historical sources of contamination.
Gas chromatographic analysis of fatty acids present in the cell membrane is the basis for a third option for microbial testing. LLI employs two MIDI Microbial Identification Systems to perform this technique, which matches the chromatographic pattern produced by esterified samples to a library of stored patterns. A new software upgrade greatly expanded the number of organisms that can be identified with the MIDI system. Because this technique is fast and cost-effective, it is often used for identifying isolates from environmental monitoring.
“With the addition of DNA sequencing, we essentially offer all of the current technologies that are used for identification, including strain typing. This gives us a broad ability to analyze a range of isolates,” explains Mark Kaiser, director of Pharmaceutical Microbiology. For more information on microbial identification, call Pharmaceutical Business Development at 717-656-2300.