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Impedance microbiology is a
microbiological Microbiology () is the scientific study of microorganisms, those being unicellular (single cell), multicellular (cell colony), or acellular (lacking cells). Microbiology encompasses numerous sub-disciplines including virology, bacteriology, ...
technique used to measure the microbial
number density The number density (symbol: ''n'' or ''ρ''N) is an intensive quantity used to describe the degree of concentration of countable objects ( particles, molecules, phonons, cells, galaxies, etc.) in physical space: three-dimensional volumetric number ...
(mainly bacteria but also
yeast Yeasts are eukaryotic, single-celled microorganisms classified as members of the fungus kingdom. The first yeast originated hundreds of millions of years ago, and at least 1,500 species are currently recognized. They are estimated to constitut ...
s) of a sample by monitoring the electrical parameters of the
growth medium A growth medium or culture medium is a solid, liquid, or semi-solid designed to support the growth of a population of microorganisms or cells via the process of cell proliferation or small plants like the moss ''Physcomitrella patens''. Different ...
. The ability of
microbial metabolism Microbial metabolism is the means by which a microbe obtains the energy and nutrients (e.g. carbon) it needs to live and reproduce. Microbes use many different types of metabolic strategies and species can often be differentiated from each other ...
to change the
electrical conductivity Electrical resistivity (also called specific electrical resistance or volume resistivity) is a fundamental property of a material that measures how strongly it resists electric current. A low resistivity indicates a material that readily allows ...
of the growth medium was discovered by Stewart and further studied by other scientists such as Oker-Blom, Parson and Allison in the first half of 20th century. However, it was only in the late 1970s that, thanks to computer-controlled systems used to monitor impedance, the technique showed its full potential, as discussed in the works of Fistenberg-Eden & Eden, Ur & Brown and Cady.


Principle of operation

When a pair of
electrode An electrode is an electrical conductor used to make contact with a nonmetallic part of a circuit (e.g. a semiconductor, an electrolyte, a vacuum or air). Electrodes are essential parts of batteries that can consist of a variety of materials de ...
s are immersed in the growth medium, the system composed of electrodes and
electrolyte An electrolyte is a medium containing ions that is electrically conducting through the movement of those ions, but not conducting electrons. This includes most soluble salts, acids, and bases dissolved in a polar solvent, such as water. Upon dis ...
can be modeled with the
electrical circuit An electrical network is an interconnection of electrical components (e.g., batteries, resistors, inductors, capacitors, switches, transistors) or a model of such an interconnection, consisting of electrical elements (e.g., voltage sources, c ...
of Fig. 1, where Rm and Cm are the resistance and
capacitance Capacitance is the capability of a material object or device to store electric charge. It is measured by the change in charge in response to a difference in electric potential, expressed as the ratio of those quantities. Commonly recognized are ...
of the bulk medium, while Ri and Ci are the resistance and capacitance of the electrode-electrolyte interface. However, when
frequency Frequency is the number of occurrences of a repeating event per unit of time. It is also occasionally referred to as ''temporal frequency'' for clarity, and is distinct from ''angular frequency''. Frequency is measured in hertz The hertz ...
of the
sinusoidal A sine wave, sinusoidal wave, or just sinusoid is a mathematical curve defined in terms of the ''sine'' trigonometric function, of which it is the graph. It is a type of continuous wave and also a smooth periodic function. It occurs often in ...
test signal applied to the electrodes is relatively low (lower than 1 MHz) the bulk capacitance Cm can be neglected and the system can be modeled with a simpler circuit consisting only of a resistance Rs and a capacitance Cs in series. The resistance Rs accounts for the electrical conductivity of the bulk medium while the capacitance Cs is due to the capacitive double-layer at the electrode-electrolyte interface. During the growth phase, bacterial metabolism transforms uncharged or weakly charged compounds of the bulk medium in highly charged compounds that change the electrical properties of the medium. This results in a decrease of resistance Rs and an increase of capacitance Cs. In impedance microbiology technique works this way, the sample with the initial unknown bacterial concentration (C0) is placed at a temperature favoring bacterial growth (in the range 37 to 42 °C if mesophilic microbial population is the target) and the electrical parameters Rs and Cs are measured at regular time intervals of few minutes by means of a couple of electrodes in direct contact with the sample. Until the bacterial concentration is lower than a critical threshold CTH the electrical parameters Rs and Cs remain essentially constant (at their baseline values). CTH depends on various parameters such as electrode geometry, bacterial strain, chemical composition of the growth medium etc., but it is always in the range 106 to 107 cfu/ml. When the bacterial concentration increases over CTH, the electrical parameters deviate from their baseline values (generally in the case of bacteria there is a decrease of Rs and an increase of Cs, the opposite happens in the case of yeasts). The time needed for the electrical parameters Rs and Cs to deviate from their baseline value is referred as Detect Time (DT) and is the parameter used to estimate the initial unknown bacterial concentration C0. In Fig. 2 a typical curve for Rs as well as the corresponding bacterial concentration are plotted vs. time. Fig. 3 shows typical Rs curves vs time for samples characterized by different bacterial concentration. Since DT is the time needed for the bacterial concentration to grow from the initial value C0 to CTH, highly contaminated samples are characterized by lower values of DT than samples with low bacterial concentration. Given C1, C2 and C3 the bacterial concentration of three samples with C1 > C2 > C3, it is DT1 < DT2 < DT3. Data from literature show how DT is a linear function of the logarithm of C0: :DT = A\cdot\log_(C_0) + B where the parameters A and B are dependent on the particular type of samples under test, the bacterial strains, the type of enriching medium used and so on. These parameters can be calculated by calibrating the system using a set of samples whose bacterial concentration is known and calculating the linear regression line that will be used to estimate the bacterial concentration from the measured DT. Impedance microbiology has different advantages on the standard plate count technique to measure bacterial concentration. It is characterized by faster response time. In the case of mesophilic bacteria, the response time range from 2 – 3 hours for highly contaminated samples (105 - 106 cfu/ml) to over 10 hours for samples with very low bacterial concentration (less than 10 cfu/ml). As a comparison, for the same bacterial strains the Plate Count technique is characterized by response times from 48 to 72 hours. Impedance microbiology is a method that can be easily automated and implemented as part of an industrial machine or realized as an embedded portable sensor, while plate count is a manual method that needs to be carried out in a laboratory by long trained personnel.


Instrumentation

Over the past decades different instruments (either laboratory built or commercially available) to measure bacterial concentration using impedance microbiology have been built. One of the best selling and well accepted instruments in the industry is the Bactometer by Biomerieux. The original instrument of 1984 features a multi-incubator system capable of monitoring up to 512 samples simultaneously with the ability to set 8 different incubation temperatures. Other instruments with performance comparable to the Bactometer are Malthus by Malthus Instruments Ltd (Bury, UK), RABIT by Don Whitley Scientific (Shipley, UK) and Bac Trac by Sy-Lab (Purkensdorf, Austria). A portable embedded system for microbial concentration measurement in liquid and semi-liquid media using impedance microbiology has been recently proposed. The system is composed of a thermoregulated incubation chamber where the sample under test is stored and a controller for thermoregulation and impedance measurements.


Applications

Impedance microbiology has been extensively used in the past decades to measure the concentration of bacteria and yeasts in different type of samples, mainly for quality assurance in the food industry. Some applications are, the determination of the shelf life of pasteurized milk and the measure of total bacterial concentration in raw-milk, frozen vegetables, grain products, meat products and beer. The technique has been also used in environmental monitoring to detect the coliform concentration in water samples as well as other bacterial pathogens like ''E.coli'' present in water bodies, in the pharmaceutical industry to test the efficiency of novel antibacterial agents and the testing of final products.


References

{{Reflist Microbiology techniques Microbiology Impedance measurements