Chemical fixation methods for Raman spectroscopy-based analysis of bacteria

Preservation of biological samples for downstream analysis is important for analytical methods that measure the biochemical composition of a sample. One such method, Raman microspectroscopy, is commonly used as a rapid phenotypic technique to measure biomolecular composition for the purposes of identification and discrimination of species and strains of bacteria, as well as investigating physiological responses to external stressors and the uptake of stable isotope-labelled substrates in single cells. This study examines the influence of a number of common chemical fixation and inactivation methods on the Raman spectrum of six species of bacteria. Modifications to the Raman-phenotype caused by fixation were compared to unfixed control samples using difference spectra and Principal Components Analysis (PCA). Additionally, the effect of fixation on the ability to accurately classify bacterial species using their Raman phenotype was determined. The results showed that common fixatives such as glutaraldehyde and ethanol cause significant changes to the Raman spectra of bacteria, whereas formaldehyde and sodium azide were better at preserving spectral features

Comparison of bacterioneuston and bacterioplankton dynamics during a phytoplankton bloom in a fjord mesocosm

The bacterioneuston is the community of Bacteria present in surface microlayers, the thin surface film that forms the interface between aquatic environments and the atmosphere. In this study we compared bacterial cell abundance and bacterial community structure of the bacterioneuston and the bacterioplankton (from the subsurface water column) during a phytoplankton bloom mesocosm experiment. Bacterial cell abundance, determined by flow cytometry, followed a typical bacterioplankton response to a phytoplankton bloom, with Synechococcus and high nucleic acid (HNA) bacterial cell numbers initially falling, probably due to selective protist grazing. Subsequently HNA and low nucleic acid (LNA) bacterial cells increased in abundance but Synechococcus did not. There was no significant difference between bacterioneuston and bacterioplankton cell abundances during the experiment. Conversely, distinct and consistent differences between the bacterioneuston and the bacterioplankton community structure were observed. This was monitored simultaneously by Bacteria 16S rRNA gene terminal restriction fragment length polymorphism (T-RFLP) and denaturing gradient gel electrophoresis (DGGE). The conserved patterns of community structure observed in all of the mesocosms indicate that the bacterioneuston is distinctive and non-random

Biotic and Abiotic Associations with Westslope Cutthroat Trout (Oncorhynchus clarkii lewisi) in the North Fork Flathead River Basin in northwestern Montana, USA and southeastern British Columbia, CAN under current and future climate scenarios.

Westslope Cutthroat Trout (Oncorhynchus clarkii lewisi; WCT) populations are declining across much of their native range due to threats such as habitat degradation, competition with non-native species, and climate change. Understanding how habitat characteristics impact distributions of nonhybridized WCT populations throughout a relatively pristine core conservation area is needed to inform management and conservation efforts. We investigated whether abiotic (e.g., gradient) and biotic (i.e., Bull Trout – Salvelinus confluentus) variables predicted WCT presence and predicted how future stream temperature projections for the area might be expected to alter distributions. We compared logistic regression models of WCT presence throughout tributaries of the North Fork Flathead River in Montana, USA and British Columbia, CAN models using a variety of metrics (e.g., Akaike Information Criterion). WCT were widespread throughout the 293 reaches analyzed (present in 69.3% of reaches). Their presence was predicted by gradient, summer temperature, and an interaction of pool density and Bull Trout. Using this regression model and climate projections under both moderate and extreme emissions scenarios, WCT presence is predicted to increase by 13.0% and 14.1% respectively in 2075 from current distributions based on changes in temperature alone. When changes in Bull Trout distributions and temperatures are considered, WCT distributions are predicted to increase by 13.4% and 17.5% under the moderate and high emissions scenario, respectively. This conservation area is predicted to continue to serve as a WCT stronghold, if other threats can be contained