Igor Buzalewicz

Influence of various growth conditions on Fresnel diffraction patterns of bacteria colonies examined in the optical system with converging spherical wave illumination

The novel optical system based on converging spherical wave illumination for analysis of bacteria colonies diffraction patterns, is proposed. The complex physical model of light transformation on bacteria colonies in this system, is presented. Fresnel diffraction patterns of bacteria colonies Escherichia coli, Salmonella enteritidis, Staphylococcus aureus grown in various conditions, were examined. It was demonstrated that the proposed system enables the characterization of morphological changes of colony structures basing on the changes of theirs Fresnel diffraction patterns.

Bacteria species identification by the statistical analysis of bacterial colonies Fresnel patterns

It was demonstrated that statistical analysis of bacteria colonies Fresnel patterns recorded in the optical system with converging spherical wave illumination is suitable for highly effective bacteria species classification. The proposed method includes Fresnel patterns recording followed by image processing and the statistical analysis based on feature extraction, feature selection, classification and classification performance methods. Examination performed on various bacteria species (Salmonella enteritidis, Staphylococcus aureus, Staphylococcus intermedius, Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa and Citrobacter freundii) revealed that the proposed method achieved very high accuracy of over 98%.

Image processing guided analysis for estimation of bacteria colonies number by means of optical transforms

A novel method for evaluation of bacterial colonies number (Colony Forming Units--CFU), is described. Proposed algorithm, based on the Mellin transform, allows the CFU evaluation, invariant for the spatial orientation and scale changes. The proposed method involves image recording of bacteria grown in Petri dishes, calculation of the Fourier spectrum followed by coordinates transformation, and determination of the Mellin transform. It was proved that there is a high correlation between CFU and maxima of Mellin spectra. The method was practically implemented for evaluation of antibacterial activity of silver-based nanomaterials and the effect of an additional laser light irradiation.

Computer-based classification of bacteria species by analysis of their colonies Fresnel diffraction patterns

In the presented paper the optical system with converging spherical wave illumination for classification of bacteria species, is proposed. It allows for compression of the observation space, observation of Fresnel patterns, diffraction pattern scaling and low level of optical aberrations, which are not possessed by other optical configurations. Obtained experimental results have shown that colonies of specific bacteria species generate unique diffraction signatures. Analysis of Fresnel diffraction patterns of bacteria colonies can be fast and reliable method for classification and recognition of bacteria species. To determine the unique features of bacteria colonies diffraction patterns the image processing analysis was proposed. Classification can be performed by analyzing the spatial structure of diffraction patterns, which can be characterized by set of concentric rings. The characteristics of such rings depends on the bacteria species. In the paper, the influence of basic features and ring partitioning number on the bacteria classification, is analyzed. It is demonstrated that Fresnel patterns can be used for classification of following species: Salmonella enteritidis, Staplyococcus aureus, Proteus mirabilis and Citrobacter freundii. Image processing is performed by free ImageJ software, for which a special macro with human interaction, was written. LDA classification, CV method, ANOVA and PCA visualizations preceded by image data extraction were conducted using the free software R.

Exploiting of optical transforms for bacteria evaluation in vitro

A novel method for evaluation of bacteria colonies concentration based on optical spectra examination, is proposed. The influence of bacteria colonies number on Fourier spectrum properties is considered in term of scalar diffraction theory and corresponding theoretical model is presented. Computational simulations are performed to confirm the theoretical predictions. Additionally, optical Mellin transform is used to omit the dependence of Fourier spectrum pattern on bacteria colonies size fluctuations and to provide a scale-invariant analysis. Presented results have shown high potential of the proposed approach for comparative study of bacteria colonies grown on solid medium in vitro.

Novel Perspectives on the Characterization of Species-Dependent Optical Signatures of Bacterial Colonies by Digital Holography

The use of light diffraction for the microbiological diagnosis of bacterial colonies was a significant breakthrough with widespread implications for the food industry and clinical practice. We previously confirmed that optical sensors for bacterial colony light diffraction can be used for bacterial identification. This paper is focused on the novel perspectives of this method based on digital in-line holography (DIH), which is able to reconstruct the amplitude and phase properties of examined objects, as well as the amplitude and phase patterns of the optical field scattered/diffracted by the bacterial colony in any chosen observation plane behind the object from single digital hologram. Analysis of the amplitude and phase patterns inside a colony revealed its unique optical properties, which are associated with the internal structure and geometry of the bacterial colony. Moreover, on a computational level, it is possible to select the desired scattered/diffracted pattern within the entire observation volume that exhibits the largest amount of unique, differentiating bacterial features. These properties distinguish this method from the already proposed sensing techniques based on light diffraction/scattering of bacterial colonies. The reconstructed diffraction patterns have a similar spatial distribution as the recorded Fresnel patterns, previously applied for bacterial identification with over 98% accuracy, but they are characterized by both intensity and phase distributions. Our results using digital holography provide new optical discriminators of bacterial species revealed in one single step in form of new optical signatures of bacterial colonies: digital holograms, reconstructed amplitude and phase patterns, as well as diffraction patterns from all observation space, which exhibit species-dependent features. To the best of our knowledge, this is the first report on bacterial colony analysis via digital holography and our study represents an innovative approach to the subject.

Bacteria identification in an optical system with optimized diffraction pattern registration condition supported by enhanced statistical analysis

It is possible to identify bacteria species basing on their diffraction patterns followed by statistical analysis. The new approach exploits two steps: optimization of the recording conditions and introduction of new interpretable features for the identification. First, optimal diffraction registration plane, was determined. Next, results were verified by the analysis workflow based on ANOVA and Fisher divergence for feature selection, QDA and SVM models for classification and identification and CV with stratified sampling, sensitivity and specificity for performance assessment of the identification process. The proposed approach resulted in high sensitivity 0.9759 and specificity 0.9903 with very small identification error 1.34%.

Degeneration of Fraunhofer diffraction on bacterial colonies due to their light focusing properties examined in the digital holographic microscope system

The degeneration of Fraunhofer diffraction conditions in the optical system with converging spherical wave illumination for bacteria species identification based on diffraction patterns is analyzed by digital holographic methods. The obtained results have shown that the colonies of analyzed bacteria species act as biological lenses with the time-dependent light focusing properties, which are characterized and monitored by means of phase retrieval from sequentially captured digital holograms. This significantly affects the location of Fraunhofer patterns observation plane, which is continuously shifted across optical axis in time.

Identification of bacteria species by using morphological and textural properties of bacterial colonies diffraction patterns

In our previous study we have shown that identification of bacteria species with the use of Fresnel diffraction patterns is possible with high accuracy and at low cost. Fresnel diffraction patterns were recorded with the optical system with converging spherical wave illumination. Obtained experimental results have shown that colonies of specific bacteria species generate unique diffraction signatures. Features used for building classification models and thus for identification were simply mean value and standard deviation calculated of pixel intensities within regions of interest called rings. This work presents new, interpretable features denoting morphological and textural properties of the Fresnel diffraction patterns and their verification with the use of the statistical analysis workflow specially developed for bacteria species identification. As data set of bacteria species diffraction patterns it is very important to find features that differentiate species in the best manner. This task includes two steps. The first is finding and extracting new, interpretable features that can potentially be better for bacteria species differentiation than the ones used before. While the second one is deciding which of them are the best for identification purposes. The new features are calculated basing on normalized diffraction patterns and central statistical moments. For the verification the analysis workflow based on ANOVA for feature selection, LDA, QDA and SVM models for classification and identification and CV, sensitivity and specificity for performance assessment of the identification process, are applied. Additionally, the Fisher divergence method also known as signal to noise ratio (SNR) for feature selection was exploited.

Diffraction signature of bacteria colonies and the influence of different incubation conditions

The optical system with converging spherical wave illumination is proposed for bacteria colony diffraction pattern analysis to distinguish bacteria species. The influence of the bacteria growth conditions on diffraction pattern is investigated.