Microbial detection

Because of minimal sample preparation and minimally invasive approaches Raman spectroscopy has the potential to gain new insights into viral and microbial biochemistry as well as a rapid identification of pathogenic microorganisms. In this chapter the unique potential of several Raman spectroscopic techniques for medical and pharmaceutical issues is discussed, which focus on the analysis of viruses as well as on microbial detection... 

There are many filter configurations within the industry, such as sheet or modular depth filter types for prefiltration purposes, flat filter membranes mainly for microbial detection and, specifications, and, most commonly, filter cartridges containing either depth filter fleeces or membrane filters. Such membrane filters are available in a large variety of membrane polymers for different applications. These materials are discussed later in the chapter. 

Phospholipids, which are responsible for the structure of the cell membrane, are easily accessed through simple extraction procedures providing a useful biomarker for microbial detection and identification. However, they are influenced by the growth conditions, nutritional status, and history of a microorganism. These factors cause changes in the phospholipid (phosphoglyceride) profile as the microbe changes its membrane composition in response to its environmental requirements. 

Fluorescence-based microbial detection systems, including epifluorescence microscopy (EFM), flow cytometry (FC) and solid-phase cytometry (SPC), lend themselves to rapid, in-situ analysis of individual microorganisms, without the need for multiplication (Lemarchand et al. 2001 Lisle et al. 2004). This review will focus on the basic principles of SPC, its advantages, disadvantages and applications, and outline some future perspectives. 

The fact that pathogenic microorganisms can be present in low numbers often hampers their detection. Conventional cnlturing may take several days and underestimate the microbial load. Therefore, more rapid and sensitive microbial detection methods wonld be nsefnl for many applications to complement or replace these traditional cnltnre methods. As SPC is a fast and sensitive tool to detect low numbers of microorganisms, it has been used for the detection and quantification of several important species. 

Chen F, Kuhlman G, Kirshner L, Matsuyama A, Sage A, Pickett M, Venkateswaran K Kem R. Sensitivity and qrplicability of a r id microbial detection system in the enumeration of bacterial spores. 104 Annual Meeting of Amaican Society for Microbiology, New Orleans, May 2004. Q4)83 page 519. 

Bioluminescence serves as an excellent reporter system as a sensitive marker for microbial detection, as a real-time, non-invasive reporter for measuring gene expression and as a measure of intracellular biochemical function (cell viability). Most widely studied of the bioluminescence systems are those belonging to the luminous bacteria Vibrio sp.. Photobacterium sp. and Photorhabdus luminescens) and the firefly Photinus pyralis). While these systems have proved extremely versatile, there are caveats to their use limiting the array of applications they can be applied to. These caveats mainly surround the nature of the luciferase enzymes, and include temperature and pH stability. 

Bioassay is the quantitative method in which the endpoint is an observable effect on a biological system or an organism. The classical approach to microbial detection involves the use of differential metabolic assays (monitored colormetrically) to determine species type in the case of most bacteria, or the use of cell culture and electron microscopy to diagnose viruses and some bacteria that are intracellular parasites. Samples taken from the environment, such as soil and water, and most clinical samples must be cultured in order to obtain sufficient numbers of various cell types for reliable identification. The time required for microbial outgrowth is typically 4-48 hours (or two weeks for certain cases, such as Mycobacterium tuberculosis). Furthermore, bacterial culture suffers from an inherent drawback cells that are viable may not be culturable, because they possess unanticipated nutritional requirements as a result of genetic mutation. 

High costs of instrumentation, maintenance, and consumables may be problematic Tests need to be developed for many relevant analytes Sample pretreatment and analyte extraction often necessary New methodologies for sample handling and processing need to be developed and optimized. This may involve use of solvents, antibody, or other extraction columns Microbial detection often requires use of growth-enrichment media before sufficient numbers are reached to allow accurate detection Some sensor systems are large, complex, and expensive... 

Aeromonas phenologenes), and lactate (Hansenula anomala). The inhibition of systems has proven successful in pyruvate monitoring Streptococcus faecium) by inhibiting glycolysis with iodoacetamide and tyrosine decarboxylase with tyramine, and in microbial detection of glutamic acid Bacillus subti-lis) treated with chloromercuribenzoate and sodium fluoride to inhibit glucose uptake and metabolism. 

There are also a number of microbial detection test kits available for use. Some of these test kits are described below. 

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