Analysis of Pathogenic Bacterial and Yeast Biofilms Using the Combination of Synchrotron ATR-FTIR Microspectroscopy and Chemometric Approaches.

Biofilms are assemblages of microbial cells, extracellular polymeric substances (EPS), and other components extracted from the environment in which they develop. Within biofilms, the spatial distribution of these components can vary. Here we present a fundamental characterization study to show differences between biofilms formed by Gram-positive methicillin-resistant Staphylococcus aureus (MRSA), Gram-negative Pseudomonas aeruginosa, and the yeast-type Candida albicans using synchrotron macro attenuated total reflectance-Fourier transform infrared (ATR-FTIR) microspectroscopy. We were able to characterise the pathogenic biofilms' heterogeneous distribution, which is challenging to do using traditional techniques. Multivariate analyses revealed that the polysaccharides area (1200-950 cm-1) accounted for the most significant variance between biofilm samples, and other spectral regions corresponding to amides, lipids, and polysaccharides all contributed to sample variation. In general, this study will advance our understanding of microbial biofilms and serve as a model for future research on how to use synchrotron source ATR-FTIR microspectroscopy to analyse their variations and spatial arrangements.

Micro- to nano-scale chemical and mechanical mapping of antimicrobial-resistant fungal biofilms.

A fungal biofilm refers to the agglomeration of fungal cells surrounded by a polymeric extracellular matrix (ECM). The ECM is composed primarily of polysaccharides that facilitate strong surface adhesion, proliferation, and cellular protection from the surrounding environment. Biofilms represent the majority of known microbial communities, are ubiquitous, and are found on a multitude of natural and synthetic surfaces. The compositions, and in-turn nanomechanical properties, of fungal biofilms remain poorly understood, because these systems are complex, composed of anisotropic cellular and extracellular material, and importantly are species and environment dependent. Therefore, genomic variation, and/or mutations, as well as environmental and growth factors can change the composition of a fungal cell's biofilm. In this work, we probe the physico-mechanical and biochemical properties of two fungal species, Candida albicans (C. albicans) and Cryptococcus neoformans (C. neoformans), as well as two antifungal resistant sub-species of C. neoformans, fluconazole-resistant C. neoformans (FlucRC. neoformans) and amphotericin B-resistant C. neoformans (AmBRC. neoformans). A new experimental methodology of characterization is proposed, employing a combination of atomic force microscopy (AFM), instrumented nanoindentation, and Synchrotron ATR-FTIR measurements. This allowed the nano-mechanical and chemical characterisation of each fungal biofilm.

Chemometrics for environmental monitoring: a review.

Environmental monitoring is necessary to ensure the overall health and conservation of an ecosystem. However, ecosystems (e.g. air, water, soil), are complex, involving numerous processes (both native and external), inputs, contaminants, and living organisms. As such, monitoring an environmental system is not a trivial task. The data obtained from natural systems is often multifaceted and convoluted, as a multitude of inputs can be intertwined within the matrix of the information obtained as part of a study. This means that trends and important results can be easily overlooked by conventional and single dimensional data analysis protocols. Recently, chemometric methods have emerged as a powerful method for maximizing the details contained within a chemical data set. Specifically, chemometrics refers to the use of mathematical and statistical analysis methods to evaluate chemical data, beyond univariant analysis. This type of analysis can provide a quantitative description of environmental measurements, while also having the capacity to reveal previously overlooked trends in data sets. Applying chemometrics to environmental data allows us to identify and describe the inter-relationship of environmental drivers, sources of contamination, and their potential impact upon the environment. This review aims to provide a detailed understanding of chemometric techniques, how they are currently used in environmental monitoring, and how these techniques can be used to improve current practices. An enhanced ability to monitor environmental conditions and to predict trends would be greatly beneficial to government and research agencies in their ability to develop environmental policies and analytical procedures.

Measuring the mechanical properties of flexible crystals using bi-modal atomic force microscopy.

Flexible crystals are an emerging class of material with unique properties and a range of potential applications. Their relatively recent development means that mechanical characterisation protocols have not yet been widely established. There is a lack of quantitative flexibility measurements, such as the elastic modulus (Young's modulus), reported in the literature. In this work, we investigate amplitude modulated-frequency modulated atomic force microscopy (AM-FM AFM) as a fast, versatile method for measuring the elastic modulus of single flexible crystals. Specifically, the elastic modulus of single crystals of copper(ii) acetylacetonate (Cu(acac)2) was measured. The elastic modulus for Cu(acac)2 was found to be 4.79 ± 0.16 GPa. Importantly, this technique was able to map the variation in mechanical properties over the surface of the material with nanoscale resolution, showing some degree of correlation between surface morphology and elastic modulus. Additionally, the distribution of elastic modulus values can be measured at different locations on the crystal, giving a statistically robust distribution, which cannot be achieved using other methods.

Imaging the air-water interface: Characterising biomimetic and natural hydrophobic surfaces using in situ atomic force microscopy.

The interface between water and a textured hydrophobic surface can exist in two regimes; either the Wenzel (surface-engulfed) or Cassie-Baxter (water-suspended) state. Better understanding of the influence of pattern geometry and spacing is crucial for the design of functional (super)hydrophobic surfaces, as inspired by numerous examples in nature. In this work, we have employed amplitude modulated - atomic force microscopy to visualize the air-water interface with an unprecedented degree of clarity on a superhydrophobic and a highly hydrophobic nanostructured surface. The images obtained provide the first real-time experimental visualization of the Cassie-Baxter wetting on the surface of biomimetic silicon nanopillars and a naturally superhydrophobic cicada wing. For both surfaces, the air-water interface was found to be remarkably well-defined, revealing a distinctly nanostructured air-water interface in the interstitial spacing. The degree of interfacial texture differed as a function of surface geometry. These results reveal that the air-water interface is heterogeneous in its structure and confirmed the presence of short-range interfacial ordering. Additionally, the overpressure values for each point on the interface were calculated, quantifying the difference in wetting behavior for the biomimetic and natural surface. Results suggest that highly-ordered, closely spaced nanofeatures facilitate robust Cassie-Baxter wetting states and therefore, can enhance the stability of (super)hydrophobic surfaces.