Label-free, non-invasive light scattering sensor for rapid screening of Bacillus colonies

Singh, Atul K.; Sun, Xiulan; Bai, Xingjian; Kim, Huisung; Abdalhaseib, Maha; Bae, Euiwon; Bhunia, Arun K.

doi:10.1016/j.mimet.2014.12.012

Cited by 23 publications

(27 citation statements)

References 67 publications

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“…We observed that the scatter patterns of the WT and mutant strains were different on BHIA and LBA but were similar on different brands of BHIA. It has been reported in our previous studies that bacterial colony scatter patterns vary with the growth medium used (5,7,9,11). Thus, we expect that any mutation in the genes involved in the biochemical pathways for substrate utilization will also result in differential scatter patterns.…”

Section: Discussionmentioning

confidence: 85%

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Virulence Gene-Associated Mutant Bacterial Colonies Generate Differentiating Two-Dimensional Laser Scatter Fingerprints

Singh

Leprun

Drolia

et al. 2016

Appl Environ Microbiol

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In this study, we investigated whether a laser scatterometer designated BARDOT (bacterial rapid detection using optical scattering technology) could be used to directly screen colonies of Listeria monocytogenes, a model pathogen, with mutations in several known virulence genes, including the genes encoding Listeria adhesion protein (LAP; lap mutant), internalin A (⌬inlA strain), and an accessory secretory protein (⌬secA2 strain). Here we show that the scatter patterns of lap mutant, ⌬inlA, and ⌬secA2 colonies were markedly different from that of the wild type (WT), with >95% positive predictive values (PPVs), whereas for the complemented mutant strains, scatter patterns were restored to that of the WT. The scatter image library successfully distinguished the lap mutant and ⌬inlA mutant strains from the WT in mixed-culture experiments, including a coinfection study using the Caco-2 cell line. Among the biophysical parameters examined, the colony height and optical density did not reveal any discernible differences between the mutant and WT strains. We also found that differential LAP expression in L. monocytogenes serotype 4b strains also affected the scatter patterns of the colonies. The results from this study suggest that BARDOT can be used to screen and enumerate mutant strains separately from the WT based on differential colony scatter patterns. IMPORTANCEIn studies of microbial pathogenesis, virulence-encoding genes are routinely disrupted by deletion or insertion to create mutant strains. Screening of mutant strains is an arduous process involving plating on selective growth media, replica plating, colony hybridization, DNA isolation, and PCR or immunoassays. We applied a noninvasive laser scatterometer to differentiate mutant bacterial colonies from WT colonies based on forward optical scatter patterns. This study demonstrates that BARDOT can be used as a novel, label-free, real-time tool to aid researchers in screening virulence gene-associated mutant colonies during microbial pathogenesis, coinfection, and genetic manipulation studies.

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Section: Discussionmentioning

confidence: 85%

“…(7), Bacillus spp. (8,9), Campylobacter spp. (10), and several genera of the Enterobacteriaceae family (11).…”

mentioning

confidence: 99%

Virulence Gene-Associated Mutant Bacterial Colonies Generate Differentiating Two-Dimensional Laser Scatter Fingerprints

Singh

Leprun

Drolia

et al. 2016

Appl Environ Microbiol

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“…Plates were screened using a prototype BARDOT machine, reported previously (Banada et al, 2009;Singh et al, 2015) and the colony scatter patterns were matched against the S. enterica scatter image libraries generated in our previous study . Initially, scatter patterns were compared with the CDC classified top 20 Salmonella serovars and non-Salmonella libraries, to determine if the isolates belonged to the genus Salmonella.…”

Section: Light Scattering Sensor-based Detectionmentioning

confidence: 99%

“…Similar reasoning was observed when libraries of Bacillus spp. containing species/strains were compared with the multiple species/strain/library (Singh et al, 2015). Scatter patterns of representative colonies of each Salmonella-positive isolate (CCRMA10, CCRMA12, CCRMA13, and CCRMA14) had PPV of 92e100% and were identified as S. enterica serovar Enteritidis after matching images with the scatter image library of top-8 and top-2 Salmonella serovars libraries.…”

Section: Detection Of Salmonella From Naturally Contaminated Food Sammentioning

confidence: 99%

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Fiber optic and light scattering sensors: Complimentary approaches to rapid detection of Salmonella enterica in food samples

et al. 2016

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Development of a multispectral light‐scatter sensor for bacterial colonies

Kim

Rajwa

Bhunia

et al. 2016

Journal of Biophotonics

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We report a multispectral elastic-light-scatter instrument that can simultaneously detect three-wavelength scatter patterns and associated optical densities from individual bacterial colonies, overcoming the limits of the single-wavelength predecessor. Absorption measurements on liquid bacterial samples revealed that the spectroscopic information can indeed contribute to sample differentiability. New optical components, including a pellicle beam splitter and an optical cage system, were utilized for robust acquisition of multispectral images. Four different genera and seven shiga toxin producing E. coli serovars were analyzed; the acquired images showed differences in scattering characteristics among the tested organisms. In addition, colony-based spectral optical-density information was also collected. The optical model, which was developed using diffraction theory, correctly predicted wavelength-related differences in scatter patterns, and was matched with the experimental results. Scatter-pattern classification was performed using pseudo-Zernike (GPZ) polynomials/moments by combining the features collected at all three wavelengths and selecting the best features via a random-forest method. The data demonstrate that the selected features provide better classification rates than the same number of features from any single wavelength. Three wavelength-merged scatter pattern from E. coli.

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Label-free, non-invasive light scattering sensor for rapid screening of Bacillus colonies

Cited by 23 publications

References 67 publications

Virulence Gene-Associated Mutant Bacterial Colonies Generate Differentiating Two-Dimensional Laser Scatter Fingerprints

Virulence Gene-Associated Mutant Bacterial Colonies Generate Differentiating Two-Dimensional Laser Scatter Fingerprints

Fiber optic and light scattering sensors: Complimentary approaches to rapid detection of Salmonella enterica in food samples

Development of a multispectral light‐scatter sensor for bacterial colonies

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