Perspective: Raman spectroscopy for detection and management of diseases affecting the nervous system.

Raman spectroscopy (RS) is used increasingly for disease detection, including diseases of the nervous system (CNS). This Perspective presents RS basics and how it has been applied to disease detection. Research that focused on using a novel Raman-based technology-Rametrix Molecular Urinalysis (RMU)-for systemic disease detection is presented, demonstrated by an example of how the RS/RMU technology could be used for detection and management of diseases of the CNS in companion animals.

Cancer detection in dogs using rapid Raman molecular urinalysis.

Introduction: The presence of cancer in dogs was detected by Raman spectroscopy of urine samples and chemometric analysis of spectroscopic data. The procedure created a multimolecular spectral fingerprint with hundreds of features related directly to the chemical composition of the urine specimen. These were then used to detect the broad presence of cancer in dog urine as well as the specific presence of lymphoma, urothelial carcinoma, osteosarcoma, and mast cell tumor.

Mitochondria Transplantation Mitigates Damage in an In Vitro Model of Renal Tubular Injury and in an Ex Vivo Model of DCD Renal Transplantation.

Objectives: To test whether mitochondrial transplantation (MITO) mitigates damage in 2 models of acute kidney injury (AKI).

Background: MITO is a process where exogenous isolated mitochondria are taken up by cells. As virtually any morbid clinical condition is characterized by mitochondrial distress, MITO may find a role as a treatment modality in numerous clinical scenarios including AKI.

Analysis of urine Raman spectra differences from patients with diabetes mellitus and renal pathologies.

Background: Chronic kidney disease (CKD) poses a major public health burden. Diabetes mellitus (DM) is one of the major causes of CKD. In patients with DM, it can be difficult to differentiate diabetic kidney disease (DKD) from other causes of glomerular damage; it should not be assumed that all DM patients with decreased eGFR and/or proteinuria have DKD.

A novel synthetic sRNA promoting protein overexpression in cell-free systems.

Bacterial small RNAs (sRNAs) that regulate gene expression have been engineered for uses in synthetic biology and metabolic engineering. Here, we designed a novel non-Hfq-dependent sRNA scaffold that uses a modifiable 20 nucleotide antisense binding region to target mRNAs selectively and influence protein expression. The system was developed for regulation of a fluorescent reporter in vivo using Escherichia coli, but the system was found to be more responsive and produced statistically significant results when applied to protein synthesis using in vitro cell-free systems (CFS).

Lambda-PCR for precise DNA assembly and modification.

Lambda-polymerase chain reaction (λ-PCR) is a novel and open-source method for DNA assembly and cloning projects. λ-PCR uses overlap extension to ultimately assemble linear and circular DNA fragments, but it allows the single-stranded DNA (ssDNA) primers of the PCR extension to first exist as double-stranded DNA (dsDNA). Having dsDNA at this step is advantageous for the stability of large insertion products, to avoid inhibitory secondary structures during direct synthesis, and to reduce costs.

Alterations in the molecular composition of COVID-19 patient urine, detected using Raman spectroscopic/computational analysis.

We developed and tested a method to detect COVID-19 disease, using urine specimens. The technology is based on Raman spectroscopy and computational analysis. It does not detect SARS-CoV-2 virus or viral components, but rather a urine 'molecular fingerprint', representing systemic metabolic, inflammatory, and immunologic reactions to infection.

Raman Spectroscopic Detection and Quantification of Macro- and Microhematuria in Human Urine.

Hematuria refers to the presence of blood in urine. Even in small amounts, it may be indicative of disease, ranging from urinary tract infection to cancer. Here, Raman spectroscopy was used to detect and quantify macro- and microhematuria in human urine samples.

Disease-Associated Multimolecular Signature in the Urine of Patients with Lyme Disease Detected Using Raman Spectroscopy and Chemometrics.

A urine-based screening technique for Lyme disease (LD) was developed in this research. The screen is based on Raman spectroscopy, iterative smoothing-splines with root error adjustment (ISREA) spectral baselining, and chemometric analysis using Rametrix software. Raman spectra of urine from 30 patients with positive serologic tests (including the US Centers for Disease Control [CDC] two-tier standard) for LD were compared against subsets of our database of urine spectra from 235 healthy human volunteers, 362 end-stage kidney disease (ESKD) patients, and 17 patients with active or remissive bladder cancer (BCA).

A synthetic biosensor to detect peroxisomal acetyl-CoA concentration for compartmentalized metabolic engineering.

Background: Sub-cellular compartmentalization is used by cells to create favorable microenvironments for various metabolic reactions. These compartments concentrate enzymes, separate competing metabolic reactions, and isolate toxic intermediates. Such advantages have been recently harnessed by metabolic engineers to improve the production of various high-value chemicals via compartmentalized metabolic engineering.

Resolving complex phenotypes with Raman spectroscopy and chemometrics.

Raman spectroscopy and chemometric analyses are used to characterize phenotypes of biological samples. The approach is relevant in biotechnology to identify and monitor productive cell cultures. It can also detect the presence of pathogens in food products and screen for disease in clinical applications.

ISREA: An Efficient Peak-Preserving Baseline Correction Algorithm for Raman Spectra.

A critical step in Raman spectroscopy is baseline correction. This procedure eliminates the background signals generated by residual Rayleigh scattering or fluorescence. Baseline correction procedures relying on asymmetric loss functions have been employed recently.

Raman chemometric urinalysis (Rametrix) as a screen for bladder cancer.

Bladder cancer (BCA) is relatively common and potentially recurrent/progressive disease. It is also costly to detect, treat, and control. Definitive diagnosis is made by examination of urine sediment, imaging, direct visualization (cystoscopy), and invasive biopsy of suspect bladder lesions.

Mitigating nutrient accumulation with microalgal growth towards enhanced nutrient removal and biomass production in an osmotic photobioreactor.

Forward osmosis (FO) has great potential for low energy consumption wastewater reuse provided there is no requirement for draw solutes (DS) regeneration. Reverse solute flux (RSF) can lead to DS build-up in the feed solution. This remains a key challenge because it can cause significant water flux reduction and lead to additional water quality problems.

A mycotoxin transporter (4D) from a library of deoxynivalenol-tolerant microorganisms.

New strategies are needed to mitigate the mycotoxin deoxynivalenol (DON) in feed and food products. Microbial DNA fragments were generated from a library of DON-tolerant microorganisms. These fragments were screened in DON-sensitive yeast strains for their ability to modify or transport DON.

Characterizing glucose, illumination, and nitrogen-deprivation phenotypes of PCC6803 with Raman spectroscopy.

Background: PCC6803 is a model cyanobacterium that has been studied widely and is considered for metabolic engineering applications. Here, Raman spectroscopy and Raman chemometrics (Rametrix™) were used to (i) study broad phenotypic changes in response to growth conditions, (ii) identify phenotypic changes associated with its circadian rhythm, and (iii) correlate individual Raman bands with biomolecules and verify these with more accepted analytical methods.

Methods: cultures were grown under various conditions, exploring dependencies on light and/or external carbon and nitrogen sources.

Characterizing metabolic stress-induced phenotypes of PCC6803 with Raman spectroscopy.

Background: During their long evolution, sp. PCC6803 developed a remarkable capacity to acclimate to diverse environmental conditions. In this study, Raman spectroscopy and Raman chemometrics tools (Rametrix) were employed to investigate the phenotypic changes in response to external stressors and correlate specific Raman bands with their corresponding biomolecules determined with widely used analytical methods.

Spectral characteristics of urine from patients with end-stage kidney disease analyzed using Raman Chemometric Urinalysis (Rametrix).

Raman Chemometric Urinalysis (RametrixTM) was used to discern differences in Raman spectra from (i) 362 urine specimens from patients receiving peritoneal dialysis (PD) therapy for end-stage kidney disease (ESKD), (ii) 395 spent dialysate specimens from those PD therapies, and (iii) 235 urine specimens from healthy human volunteers. RametrixTM analysis includes spectral processing (e.g.

Dipstick analysis of urine chemistry: benefits and limitations of dry chemistry-based assays.

Urinalysis is a commonly utilized laboratory test, and analysis of urine has been studied and used since ancient times. Urine contains a wide array of metabolites that can provide information regarding the current physiologic state of the body and clinical manifestations of disease. In this review, we discuss the mechanics of the dry chemistry component of the urine dipstick such as the reaction principles underlying various assays and potential effects of collection and storage on results.

Spectral characteristics of urine specimens from healthy human volunteers analyzed using Raman chemometric urinalysis (Rametrix).

Raman chemometric urinalysis (Rametrix™) was used to analyze 235 urine specimens from healthy individuals. The purpose of this study was to establish the "range of normal" for Raman spectra of urine specimens from healthy individuals. Ultimately, spectra falling outside of this range will be correlated with kidney and urinary tract disease.

Identification of soil bacteria capable of utilizing a corn ethanol fermentation byproduct.

A commercial corn ethanol production byproduct (syrup) was used as a bacterial growth medium with the long-term aim to repurpose the resulting microbial biomass as a protein supplement in aquaculture feeds. Anaerobic batch reactors were used to enrich for soil bacteria metabolizing the syrup as the sole nutrient source over an eight-day period with the goal of obtaining pure cultures of facultative organisms from the reactors. Amplification of the V4 variable region of the 16S rRNA gene was performed using barcoded primers to track the succession of microbes enriched for during growth on the syrup.

3D printed conformal microfluidics for isolation and profiling of biomarkers from whole organs.

The ability to interface microfluidic devices with native complex biological architectures, such as whole organs, has the potential to shift the paradigm for the study and analysis of biological tissue. Here, we show 3D printing can be used to fabricate bio-inspired conformal microfluidic devices that directly interface with the surface of whole organs. Structured-light scanning techniques enabled the 3D topographical matching of microfluidic device geometry to porcine kidney anatomy.

Modification of the Mycotoxin Deoxynivalenol Using Microorganisms Isolated from Environmental Samples.

The trichothecene mycotoxin deoxynivalenol (DON) is a common contaminant of wheat, barley, and maize. New strategies are needed to reduce or eliminate DON in feed and food products. Microorganisms from plant and soil samples collected in Blacksburg, VA, USA, were screened by incubation in a mineral salt media containing 100 μg/mL DON and analysis by gas chromatography mass spectrometry (GC/MS).

Characterization of an evolved carotenoids hyper-producer of Saccharomyces cerevisiae through bioreactor parameter optimization and Raman spectroscopy.

An evolutionary engineering approach for enhancing heterologous carotenoids production in an engineered Saccharomyces cerevisiae strain was used previously to isolate several carotenoids hyper-producers from the evolved populations. β-Carotene production was characterized in the parental and one of the evolved carotenoids hyper-producers (SM14) using bench-top bioreactors to assess the impact of pH, aeration, and media composition on β-carotene production levels. The results show that with maintaining a low pH and increasing the carbon-to-nitrogen ratio (C:N) from 8.