Unlike last year, the past 12 months for microbiology were not influenced by influenza, unique strains associated with specific disease entities or emergence of multi-drug resistant isolates from around the world. In fact, 2010 seemed to be more of a stable year regarding laboratory methodologies.
Although not inclusive, below are several topics that will influence microbiology, laboratory medicine and public health in the next 3 to 5 years.
New Molecular Approaches
Up to this time the announcement of molecular methods and PCR have not reached the anticipated impact in microbiology or in other rapid methods of detection and recognition. Molecular methods have been applied mostly to virology, although the application in some areas for screening have had impact, particularly methicillin resistant Staphylococcus.
Newer options are becoming available that will have a dramatic impact in microbiology. Direct sequencing from patient samples will address rapid detection of endocarditis, bone and joint infection. The most common target will be the 16-SRNA gene or other liposomal genes and will likely be linked to sterile sites where there is no endogenous flora in the identification of one organism. The problem of mixed infections is always an issue.
As we begin to expand our understanding of microbial and endogenous flora, detection of microbial populations and the diversity of such may be the answer rather than specific identification of individual organisms.
This will probably focus on T-RFLP profiling. It, too, is focusing on 16-SRNA, but the mixture of amplicons is subject to restriction enzyme digestion and the mixture of fragments separated by electrophoresis.
The TRFLP profiling has had significant impact in environmental samples, oral flora and environmental sampling best associated with multiple rather than single species.
Also called deep sequencing or high throughput sequencing, next-generation sequencing (NGS) has the amplification of genetic material by PCR and uses ligation of amplified material to a solid surface. Sequencing is done in a massive parallel fashion; sequence information is captured by unique software.
This methodology will be particularly applicable to the pathologic process in patients with non-detectable organisms or viable but non-culturable. The most unique of the NGS is now called 454, which uses the principals of pyrosequencing utilizing detection of pyrophosphate release upon nucleotide incorporation. Advantages of NGS are that there is no bacterial cloning. It is faster, less labor-intensive, more cost effective and has a higher sensitivity than array-based detection. It is particularly suitable for pathogen discovery. However, it also is quite expensive.
Protein Mass Spectrometry
Given its rapid analysis at a relatively inexpensive cost per assay, protein mass spectrometry (PMS) may be the future of microbiology. It utilizes three functional units--ionization source, analyzer and detector. The analyzer separates ions according to mass-to-charge ratio and the detector separates ions and identifies their relative abundance.
MALDI-TOF (matrix-assisted laser disorption ionization--time of flight) is the most recognized of the PMS systems on the market. The Brooker Dalton ICS MALDI BIO-TYPERTM has been used extensively in Europe with more than 250 sites reporting significant time savings and economy of testing. BD also has MALDI-type products in the pipeline.
The benefit of these systems is not only the system, but the reality that there is an established profile and matrix with each organism determined by the manufacturers and allows for the identification within seconds or minutes of sample received in the laboratory. Generally, the MALDI-TOF offers two to four hours in complete turnaround of a specimen and the organism recovered on a detectable culture media.
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