Abstract
Label-free bacterial colony phenotyping technology called BARDOT (Bacterial Rapid Detection using Optical scattering Technology) provided successful classification of several different bacteria at the genus, species, and serovar level. Recent experiments with colonies of Bacillus species provided strikingly different characteristics of elastic light scatter (ELS) patterns, which were comprised of random speckles compared to other bacteria, which are dominated by concentric rings and spokes. Since this laser-based optical sensor interrogates the whole volume of the colony, 3-D information of micro- and macro-structures are all encoded in the far-field scatter patterns. Here, we present a theoretical model explaining the underlying mechanism of the speckle formation by the colonies from Bacillus species. Except for Bacillus polymyxa, all Bacillus spp. produced random bright spots on the imaging plane, which presumably dependent on the cellular and molecular organization and content within the colony. Our scatter model-based analysis revealed that colony spread resulting in variable surface roughness can modify the wavefront of the scatter field. As the center diameter of the Bacillus spp. colony grew from 500 to 900 μm, average speckles area decreased two-fold and the number of small speckles increased seven-fold. In conclusion, as Bacillus colony grows, the average speckle size in the scatter pattern decreases and the number of smaller speckle increases due to the swarming growth characteristics of bacteria within the colony.
Highlights
Bacterial colonies consist of millions of individual cells interacting with each other in complex ways (Shapiro, 1992; Wimpenny, 1992; Shimada et al, 1995; Ben-Jacob et al, 1998)
We investigated the different optical properties of these two Bacillus species; (i) to understand how B. subtilis and B. polymyxa construct their colony using recently developed Integrated Colony Morphology Analyzer (ICMA) (Kim et al, 2013); (ii) to compare the theoretical prediction of elastic light scatter (ELS) patterns with experimental data; and (iii) to calculate the speckle statistics to quantitatively correlate the optical phase modulation into the structure of the bacterial colony
FORWARD SCATTEROMETER Scatterometer (Figure 1A), consists of laser diode (LD) with 635 nm wavelength that was installed as a light source, and the light source directly illuminates a single bacterial colony grown on semi-solid agar plate
Summary
Bacterial colonies consist of millions of individual cells interacting with each other in complex ways (Shapiro, 1992; Wimpenny, 1992; Shimada et al, 1995; Ben-Jacob et al, 1998). Shapiro and Wimpenny first observed bacterial colony formation in time and space (Shapiro, 1992; Wimpenny, 1992) and since many other researchers employed both experimental (Shimada et al, 1995; Ben-Jacob et al, 1998; Bees et al, 2000; Stecchini et al, 2001; Kaito and Sekimizu, 2007; Pipe and Grimson, 2008) and theoretical models (Kawasaki et al, 1997; Cohen et al, 1999; Kozlovsky et al, 1999; Lega and Passot, 2003) to explain such phenomenon. We studied Bacillus as a model organism to correlate the swarming colony morphology to the optical light scattering and speckle effect
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