Mid-infrared spectroscopic assessment of nanotoxicity in Gram-negative vs. Gram-positive bacteria

Abstract

Vibrational spectroscopy provides a spectral fingerprint identifying the effects of carbonbased nanoparticles in bacterial cells. 3 ABSTRACTNanoparticles appear to induce toxic effects through a variety of mechanisms including generation of reactive oxygen species (ROS), physical contact with the cell membrane and indirect catalysis due to remnants from manufacture. The development and subsequent increasing usage of nanomaterials has highlighted a growing need to characterize and assess the toxicity of nanoparticles, particularly those that may have detrimental health effects such as carbon-based nanomaterials (CBNs). Due to interactions of nanoparticles with some reagents, many traditional toxicity tests are unsuitable for use with CBNs. Infrared (IR) spectroscopy is a non-destructive, high throughput technique, which is unhindered by such problems. We explored the application of IR spectroscopy to investigate the effects of CBNs on Gram-negative (Pseudomonas fluorescens) and Gram-positive (Mycobacterium vanbaalenii PYR-1) bacteria. Two types of IR spectroscopy were compared: attenuated total reflection Fourier-transform infrared (ATR-FTIR) and synchrotron radiation-based FTIR (SR-FTIR) spectroscopy. This showed that Gram-positive and Gram-negative bacteria exhibit differing alterations when exposed to CBNs. Gram-positive bacteria appear more resistant to these agents and this may be due to the protection afforded by their more sturdy cell wall. Markers of exposure also vary according to Gram status; Amide II was consistently altered in Gram-negative bacteria and carbohydrate altered in Gram-positive bacteria. ATR-FTIR and SR-FTIR spectroscopy could both be applied to extract biochemical alterations induced by each CBN that were consistent across the two bacterial species; these may represent potential biomarkers of nanoparticle-induced alterations. Vibrational spectroscopy approaches may provide a novel means of fingerprinting the effects of CBNs in target cells.

Fingerprinting microbiomes towards screening for microbial antibiotic resistance

“…Riding et al applied IR spectroscopy to fingerprint microbes following exposure to carbon nanoparticles, and revealed concentration-and size-dependent changes in cellular components 104 . Another study 105 confirmed the ability of IR spectroscopy to characterize changes induced by carbon nanoparticles via investigating their effects in both Gram positive and negative bacteria. Some discriminant biochemical markers, i.e., Amide II and carbohydrate, were picked out to verify the distinct alterations in bacteria with respective cell wall structures.…”

“…Detailed information of the most widely used peak frequencies and their assignments refers to the reviews of IR spectroscopy 108 , Raman spectroscopy 52,87 , and ratios (X cm -1 /Y cm -1 ) 109 . These peaks with remarkable alterations are determined as discriminating biomarkers for diagnosis of changes resulted by specific exposures in many studies 69,87,104,105,108,110 . However, no biospectroscopy-relevant study has yet been found associated with biomarkers for antibiotic 13 resistance of microbiomes.…”

Spectrochemical analyses of growth phase-related bacterial responses to low (environmentally-relevant) concentrations of tetracycline and nanoparticulate silver

“…730785, Sigma-Aldrich, UK) have a 10-nm particle size at a stock concentration of 0.02 mg/mL, dissolved in aqueous buffer with sodium citrate as stabilizer. Two classic soil bacterial strains, Mycobacterium vanbaalenii PYR-1 (Gram-positive) and Pseudomonas fluorescens (Gram-negative), were selected owing to their ubiquity and wide distribution in the soil environment and their well-known physiological behaviour 36,[46][47][48] . These two strains were cultured in Luria-Bertani (LB) broth at 30±2 o C with 150 rpm shaking for 24 h. Bacterial growth was measured every 10 min by optical density at 600 nm (OD 600 ) with a microplate reader (FLUOstar Omega, BMG Labtech, UK).…”

“…Specifically, infrared (IR) spectroscopy exploits the principle that biochemical bonds perform some degree of vibrations induced by stretching, bending, scissoring or twisting after energy absorption at particular wavelengths. The "biochemical-cell fingerprint" region is located within the mid-IR region, which is the most information-rich about biochemical structures 36 . Through assessing derived spectral peaks or alterations, i.e., bio-fingerprints, the biochemical structure of interrogated targets can be revealed [37][38][39] .…”

Diet-sourced carbon-based nanoparticles induce lipid alterations in tissues of zebrafish (Danio rerio) with genomic hypermethylation changes in brain

“…Applying a computational framework to a generated data set allows one to extract distinguishing features associated with altered chemical bonds; these can be associated with constituents such as lipids, proteins or DNA/RNA (24). Raman microspectroscopy has been applied to shed insights into CBN-induced effects (25,26), cell lineage (27) and to generate tissue images (28). …”