Metabolomic Profiling of Nutrient Solutions for Characterization of Human Gut Microbiota

Studying the metabolite output, the “exometabolome,” of bacterial communities provides insight into the metabolic interactions occurring within that ecosystem. As such, a metabolomic approach can be used to gain further understanding of the human gut microbiota. As the microbes that reside in the human gastrointestinal (GI) system, the ecosystem has extensive impacts on the human body. In vitro systems for culturing gut microbes circumvents the ethical and technical challenges of in vivo studies, while, at the same time, lends itself to metabolomic investigations. The nutrient solutions in which microbial communities representative of the human gut microbiota are cultured contain a wealth of metabolomic data. In this thesis, metabolomic characterization is used to validate an in vitro gut-mimicking bioreactor system. This provided support for the ability to establish standardized protocols on preparing, culturing and reporting on in vitro gut microbiota. Furthermore, the metabolomic approach to studying gut microbes was used in applications relevant to clinical and industrial research. The findings from this thesis demonstrate the application of a metabolomic approach to studying the human gut microbiota in a single-stage continuous stirred tank reactor (CSTR) system. More importantly, the research here exemplifies the ways in which metabolomic insight can be translated into deciphering the complex interactions of the human gut microbiota

Corrigendum to “Effects of therapeutic hypothermia on the gut microbiota and metabolome of infants suffering hypoxic-ischemic encephalopathy at birth” [Int. J. Biochem. Cell Biol. 93 (December) (2017), 110-118]

peer-reviewedCorrigendum Refers to: Watkins, C., Murphy, K., Yen, S., Carafa, I., Dempsey, E., O’Shea, C., Vercoe, E., Ross, R., Stanton, C. and Ryan, C. (2017). Effects of therapeutic hypothermia on the gut microbiota and metabolome of infants suffering hypoxic-ischemic encephalopathy at birth. The International Journal of Biochemistry & Cell Biology, [online] 93, pp.110-118. Available at: https://doi.org/10.1016/j.biocel.2017.08.01

isolateR: an R package for generating microbial libraries from Sanger sequencing data

Motivation: Sanger sequencing of taxonomic marker genes (e.g. 16S/18S/ITS/rpoB/cpn60) represents the leading method for identifying a wide range of microorganisms including bacteria, archaea, and fungi. However, the manual processing of sequence data and limitations associated with conventional BLAST searches impede the efficient generation of strain libraries essential for cataloging microbial diversity and discovering novel species. Results: isolateR addresses these challenges by implementing a standardized and scalable three-step pipeline that includes: (1) automated batch processing of Sanger sequence files, (2) taxonomic classification via global alignment to type strain databases in accordance with the latest international nomenclature standards, and (3) straightforward creation of strain libraries and handling of clonal isolates, with the ability to set customizable sequence dereplication thresholds and combine data from multiple sequencing runs into a single library. The tool’s user-friendly design also features interactive HTML outputs that simplify data exploration and analysis. Additionally, in silico benchmarking done on two comprehensive human gut genome catalogues (IMGG and Hadza hunter-gather populations) showcase the proficiency of isolateR in uncovering and cataloging the nuanced spectrum of microbial diversity, advocating for a more targeted and granular exploration within individual hosts to achieve the highest strain-level resolution possible when generating culture collections. Availability and implementation: isolateR is available at: https://github.com/bdaisley/isolateR

Treating Cell Culture Media with UV Irradiation against Adventitious Agents: Minimal Impact on CHO Performance

Sterility of cell culture media is an important concern in biotherapeutic processing. In large scale biotherapeutic production, a unit contamination of cell culture media can have costly effects. Ultraviolet (UV) irradiation is a sterilization method effective against bacteria and viruses while being non-thermal and non-adulterating in its mechanism of action. This makes UV irradiation attractive for use in sterilization of cell culture media. The objective of this study was to evaluate the effect of UV irradiation of cell culture media in terms of chemical composition and the ability to grow cell cultures in the treated media. The results showed that UV irradiation of commercial cell culture media at relevant disinfection doses impacted the chemical composition of the media with respect to several carboxylic acids, and to a minimal extent, amino acids. The cumulative effect of these changes, however, did not negatively influence the ability to culture Chinese Hamster Ovary cells, as evaluated by cell viability, growth rate, and protein titer measurements in simple batch growth compared with the same cells cultured in control media exposed to visible light. (C) 2014 American Institute of Chemical EngineersNSERC ENGAGE; NSERC Strategic Network (MabNet); NSERC Canada Graduate Scholarshi

Inexpensive method for producing macroporous silicon particulates (MPSPs) with pyrolyzed polyacrylonitrile for lithium ion batteries

One of the most exciting areas in lithium ion batteries is engineering structured silicon anodes. These new materials promise to lead the next generation of batteries with significantly higher reversible charge capacity than current technologies. One drawback of these materials is that their production involves costly processing steps, limiting their application in commercial lithium ion batteries. In this report we present an inexpensive method for synthesizing macroporous silicon particulates (MPSPs). After being mixed with polyacrylonitrile (PAN) and pyrolyzed, MPSPs can alloy with lithium, resulting in capacities of 1000 mAhg−1 for over 600+ cycles. These sponge-like MPSPs with pyrolyzed PAN (PPAN) can accommodate the large volume expansion associated with silicon lithiation. This performance combined with low cost processing yields a competitive anode material that will have an immediate and direct application in lithium ion batteries

Perceived racial/ethnic discrimination, smoking and alcohol consumption in the Multi-Ethnic Study of Atherosclerosis (MESA)

OBJECTIVE: To examine the association of perceived racial/ethnic discrimination with smoking and alcohol consumption in adults participating in the Multi-Ethnic Study of Atherosclerosis. METHODS: Data on 6680 black, Chinese, Hispanic and white adults aged 45 to 84 years of age recruited from Illinois, New York, Maryland, North Carolina, Minnesota and California during 2000 and 2002 were used for this analysis. Logistic regression was used to estimate the association of perceived racial/ethnic discrimination with smoking status and alcohol consumption for each racial/ethnic group separately. RESULTS: Blacks were more likely to experience racial/ethnic discrimination (43%) than Hispanics (19%), Chinese participants (10%) or whites (4%, P<0.0001). In the fully-adjusted model, blacks reporting racial/ethnic discrimination had 34% and 51% greater odds of reporting smoking and drinking, respectively, than blacks who did not report racial/ethnic discrimination. Hispanics reporting racial/ethnic discrimination had 62% greater odds of heavy drinking. Whites reporting racial/ethnic discrimination had 88% greater odds of reporting being current smokers than whites who did not report racial/ethnic discrimination. CONCLUSIONS: Our findings suggest that the experience of discrimination is associated with greater prevalence of unhealthy behaviors. Specifically, the use of smoking and alcohol may be patterned by experience of discrimination. PMID: 20609433 [PubMed - in process]PMCID: PMC2939242 [Available on 2011/9/1]Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/78013/3/2010BorellLNPerceivedracialethnicdiscrimination.pd

The effect of a hydrolyzed protein diet on the fecal microbiota in cats with chronic enteropathy

The effect of a hydrolyzed protein diet on the fecal microbiota has not been studied in feline chronic enteropathy (CE). Our study aimed to (1) compare the fecal microbiota of cats with CE to control cats with no gastrointestinal signs and (2) determine the effect of a hydrolyzed protein diet on the fecal microbiota of cats with CE and whether this differs between dietary responders and non-responders. The fecal microbiome of cats with CE (n = 36) showed decreased α-diversity in terms of genus richness (P = 0.04) and increased β-diversity in terms of Bray-Curtis Dissimilarity (P &lt; 0.001) compared to control cats (n = 14). Clostridium was the only genera significantly over-represented in cats with CE compared to control cats (adjusted P &lt; 0.1). After 6-weeks of feeding the diet, fifteen cats were classified as responders and 18 as non-responders, based on clinical signs. At the genus level, α-diversity was increased in non-responders versus responders at diagnosis, but decreased after dietary intervention in both groups (P &lt; 0.05). At the family level, non-responders became increasingly dissimilar after dietary intervention (P = 0.012). In general, the abundance of bacteria decreased with feeding a hydrolyzed diet, with the genera most significantly affected being more frequently observed in non-responders. Bifidobacterium was the only genus that increased significantly in abundance post-diet and this effect was observed in both responders and non-responders. Both Oscillibacter and Desulfovibrionaceae_unclassified were most abundant in non-responders at diagnosis but were rarely observed post diet in neither responders nor non-responders. Cats with CE had similar microbiota changes to those described in human inflammatory bowel disease. Whether the presence of Oscillibacter and Desulfovibrionaceae_unclassified are indicators of non-response to the diet at diagnosis requires further investigation. Despite the hydrolyzed diet reducing α-diversity in all cats with CE, this did not resolve gastrointestinal signs in some cats. However, responders metabolized the diet in a similar manner, reflected by sustained β-diversity, while the microbiome of non-responders became increasingly dissimilar compared to diagnosis at the family level. Therefore, the microbiome may not be as tightly regulated in cats with CE that are non-responders and therefore, these cats would require additional therapy for remission of clinical signs

The effects of a hydrolyzed protein diet on the plasma, fecal and urine metabolome in cats with chronic enteropathy

Hydrolyzed protein diets are extensively used to treat chronic enteropathy (CE) in cats. However, the biochemical effects of such a diet on feline CE have not been characterized. In this study an untargeted 1H nuclear magnetic resonance spectroscopy-based metabolomic approach was used to compare the urinary, plasma, and fecal metabolic phenotypes of cats with CE to control cats with no gastrointestinal signs recruited at the Royal Veterinary College (RVC). In addition, the biomolecular consequences of a hydrolyzed protein diet in cats with CE was also separately determined in cats recruited from the RVC (n = 16) and the University of Bristol (n = 24) and whether these responses differed between dietary responders and non-responders. Here, plasma metabolites related to energy and amino acid metabolism significantly varied between CE and control cats in the RVC cohort. The hydrolyzed protein diet modulated the urinary metabolome of cats with CE (p = 0.005) in both the RVC and Bristol cohort. In the RVC cohort, the urinary excretion of phenylacetylglutamine, p-cresyl-sulfate, creatinine and taurine at diagnosis was predictive of dietary response (p = 0.025) although this was not observed in the Bristol cohort. Conversely, in the Bristol cohort plasma betaine, glycerol, glutamine and alanine at diagnosis was predictive of outcome (p = 0.001), but these same results were not observed in the RVC cohort. The biochemical signature of feline CE in the RVC cohort was consistent with that identified in human and animal models of inflammatory bowel disease. The hydrolyzed protein diet had the same effect on the urinary metabolome of cats with CE at both sites. However, biomarkers that were predictive of dietary response at diagnosis differed between the 2 sites. This may be due to differences in disease severity, disease heterogeneity, factors unrelated to the disease or small sample size at both sites. As such, further studies utilizing larger number of cats are needed to corroborate these findings.</p