Metagenomics Approaches in Advancing Microbiome Research

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Description
Metagenomics is the study of the structure and function of microbial communities through the application of the whole-genome shotgun (WGS) sequencing method. Providing high-resolution community profiles at species or even strain levels, metagenomics points to a new direction for microbiome…

Metagenomics is the study of the structure and function of microbial communities through the application of the whole-genome shotgun (WGS) sequencing method. Providing high-resolution community profiles at species or even strain levels, metagenomics points to a new direction for microbiome research in understanding microbial gene function, microbial-microbial interactions, and host-microbe interactions. My thesis work includes innovation in metagenomic research through the application of ChatGPT in assisting beginning researchers, adopt pre-existed alpha diversity metric for metagenomic data to improve diversity calculation, and the application of metagenomic data in Alzheimer’s disease research.Since the release of ChatGPT in March 2023, the conversation regarding AI in research has promptly been debated. Through the prompted bioinformatic case study, I demonstrate the application of ChatGPT in conducting metagenomic analysis. I constructed and tested a working pipeline aimed at instructing GPT in completing shotgun metagenomic research. The pipeline includes instructions for various essential analytic steps: quality controls, host filtering, read classification, abundance estimation, diversity calculation, and data visualization. The pipeline demonstrated successful completion and reproducible results. Alpha diversity measurement is critical to understanding microbiomes. The widely used Faith’s phylogenetic diversity (PD) metric is agnostic of feature abundance and, therefore, falls short of analyzing metagenomic data. BWPDθ, an abundance weighted variant of Faith’s PD, was implemented in scikit-bio alpha diversity metrics. My analysis shows that BWPDθ does have better performance compared to Faith’s PD, revealing more biological significance, and maintaining their robustness at a lower sampling depth. The progression of Alzheimer’s disease (AD) is known to be associated with alterations in the patient’s gut microbiome. Utilizing metagenomic data from the AlzBiom study, I explored the differential abundance of bacterial pncA genes among healthy and AD participants by age group. The analysis showed that there was no significant difference in pncA abundance between the healthy and AD patients. However, when stratified by age group, within the age group 64 to 69, AD was shown to have significantly lower pncA abundance than the healthy control group. The Pearson's test showed a moderate positive association between age and pncA abundance.
Date Created
2024
Agent

Halogenases Involved in Complex Biosynthesis

Description
With uses in fields such as medicine, agriculture, and biotechnology, halogenases are useful enzymes in nature which add or substitute halogens onto other molecules. By doing so, they become necessary for biosynthesis and cross-coupling reactions. Halogenases can be classified by…

With uses in fields such as medicine, agriculture, and biotechnology, halogenases are useful enzymes in nature which add or substitute halogens onto other molecules. By doing so, they become necessary for biosynthesis and cross-coupling reactions. Halogenases can be classified by three main types of mechanisms: nucleophilic, radical, and electrophilic. From there, they can be further broken down by the halogen involved, the substrate needed, other proteins used, or molecules generated. A notable example is PrnA which is a tryptophan-7 halogenase that falls under the flavin-dependent definition with an electrophilic mechanism. Historically, research on these enzymes was slow until the use of bioinformatics rapidly accelerated discoveries to the point where halogenases like VirX1 can be identified from viruses. By reviewing the literature available on halogenase since their first analysis, a better understanding of their functions can be obtained. Also, with the application of bioinformatics, a phylogenetic analysis on the halogenases present in cyanobacteria can be conducted and compared.
Date Created
2024-05
Agent

The Effect of Hydration Pulse Duration on the Diversity and Composition of Soil Microbiomes

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Description
The study of organismal adaptations oftentimes focuses on specific, constant conditions, but environmental parameters are characterized by more or less marked levels of variability, rather than constancy. This is important in environments like soils where microbial activity follows pulses of…

The study of organismal adaptations oftentimes focuses on specific, constant conditions, but environmental parameters are characterized by more or less marked levels of variability, rather than constancy. This is important in environments like soils where microbial activity follows pulses of water availability driven by precipitation. Nowhere are these pulses more variable and unpredictable than in arid soils. Pulses constitute stressful conditions for bacteria because they cause direct cellular damage that must be repaired and they force cells to toggle between dormancy and active physiological states, which is energetically taxing. I hypothesize that arid soil microorganisms are adapted to the variability in wet/dry cycles itself, as determined by the frequency and duration of hydration pulses. To test this, I subjected soil microbiomes from the Chihuahuan Desert to controlled incubations for a total common growth period of 60 hours, but separated into treatments in which the total active time was reached with hydration pulses of different length with intervening periods of desiccation, so as to isolate pulse length and frequency as the varying factors in the experiment. Using 16S rRNA amplicon data, I characterized changes in microbiome growth, diversity, and species composition, and tracked the individual responses to treatment intensity in the 447 most common bacterial species (phylotypes) in the soil. Considering knowledge of extremophile microbiology, I hypothesized that growth yield and diversity would decline with shorter pulses. I found that microbial diversity was indeed a direct function of pulse length, but surprisingly, total yield was an inverse function of it. Pulse regime treatments resulted in progressively more significant differences in community composition with increasing pulse length, as differently adapted phylotypes became more prominent. In fact, more than 30% of the most common bacterial phylotypes demonstrated statistically significant population growth responses to pulse length. Most responsive phylotypes were apparently best adapted to short pulse regimes (known in the literature as Nimble Responders or NIRs), while fewer did better under long pulse regimes (known as TORs or Torpid Responders). Examples of extreme NIRs and TORs could be found among bacteria from different phyla, indicating that these adaptations have occurred multiple times during evolution.
Date Created
2023
Agent

Discovering the Association between Gut Microbiome and Occurrence of Chemotherapy-Induced Nausea Among Breast Cancer Patients

Description

Humans and their microbiota are in a symbiotic relationship. It is known that microbiale residents within and on human bodies have the potential to impact host physiology in both healthy and disease states. To date, little is known about the…

Humans and their microbiota are in a symbiotic relationship. It is known that microbiale residents within and on human bodies have the potential to impact host physiology in both healthy and disease states. To date, little is known about the potential influence of the gut microbiome on the onset of nausea symptoms among cancer patients undergoing chemotherapy treatment. Chemotherapy-induced nausea (CIN) is a serious and common side effect. The CIN presentation is often coupled with other symptoms such as fatigue, sleep disturbance, depression, and anxiety. These symptoms both on an individual and collective level, cause negative impacts on patients’ health outcome as they challenge patients’ ability to tolerate and comply with chemotherapy. To understand the association between gut microbiome and CIN, we applied 16S rRNA amplicon sequencing to characterize the gut microbiome of breast cancer patients who reported nausea symptoms and those who reported no nausea symptoms. We hypothesize that the gut microbiome of patients who reported nausea symptoms is distinct from patients who reported no nausea. Our findings support this hypothesis, as the gut microbiome of nausea case was distinct from the no nausea cases. Specifically, we observed decreased abundance of Bacteroidetes in patients who reported nausea, while patients who reported no CIN had constant or increased abundance of Bacteroidetes. Overall, we showed that changes in the gut microbiota have an association with the occurrence of CIN symptoms among breast cancer patients receiving chemotherapy. These findings provide preliminary data for extensive research on the role of gut microbiome in CIN in the future.

Date Created
2023-05
Agent

Deciphering Connections Between the Microbial Communities of the Euphotic Zone and Sinking Particles in the Sargasso Sea

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Description
Sinking particles are important conduits of organic carbon from the euphotic zone to the deep ocean and microhabitats for diverse microbial communities, but little is known about what determines their origin and community composition. Events in the northwestern Sargasso Sea,…

Sinking particles are important conduits of organic carbon from the euphotic zone to the deep ocean and microhabitats for diverse microbial communities, but little is known about what determines their origin and community composition. Events in the northwestern Sargasso Sea, such as winter convective mixing, summer stratification, and mesoscale (10–100 km) eddies, characteristic features of this region, affect the vertical and temporal composition and abundance of pelagic and particle-attached microorganisms. To assess the connections of the microbial communities between the euphotic zone and sinking particles, I carried out indicator and differential abundance analyses of prokaryotes and photoautotrophs based on the V4-V5 amplicons of the 16S rDNA from samples collected in the Sargasso Sea during the spring and summer of 2012. I found that gammaproteobacteria such as Pseudoalteromonas sp. and Vibrio sp., common particle-associated bacteria often linked with zooplankton, dominated the sequence libraries of the sinking particles. The analysis also revealed that members of Flavobacteria, particularly the fish pathogen Tenacibaculum sp., as well as Chloropicon sp. and Chloroparvula sp., among the smallest known green algae, were indicators taxa of sinking particles. The cryptophyte Teleaulax and the diatom Chaetoceros were overrepresented in the particle communities during both seasons. Interestingly, I also found that the large centric diatom, Rhizosolenia sp., generally rare in the oligotrophic Sargasso Sea, dominated photoautotrophic communities of sinking particles collected in the center of an anticyclonic eddy with unusual upwelling due to eddy-wind interactions. I hypothesize that the steady contribution by picophytoplankton to particle flux is punctuated by pulses of production and flux of larger-sized phytoplankton in response to episodic eddy upwelling events and can lead to higher export of particulate organic matter during the summer.
Date Created
2022
Agent

Interactions between the Gut Microbiome and Serp-1 after Acute Renal Rejection

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Description
Current data indicates that between 10-20% of patients that receive renal transplants experience acute rejection. Acute rejection is an important predictor of future complications that can lead to complete graft loss. Serine Protease Inhibitor proteins (Serpins) have shown incredible promise…

Current data indicates that between 10-20% of patients that receive renal transplants experience acute rejection. Acute rejection is an important predictor of future complications that can lead to complete graft loss. Serine Protease Inhibitor proteins (Serpins) have shown incredible promise in working to lessen or inhibit the onset and effects of acute rejection in multiple solid organ transplant studies. However, it has been proposed that the mechanisms of Serp-1 immunomodulation can be affected by the native gut microbiome of recipient animals. This study utilizes murine renal allograft transplant from C57BL/6 to BALB/c and microbiome depletion to model the acute rejection response and to determine if there are interactions between the gut microbiome and Serp- 1 during the acute renal allograft rejection response. After antibiotic administration, expected decreases in richness and diversity in the designated group were observed. After renal subcapsular allograft transplant, histological and microbiome analyses revealed neither significant microbiome changes persistent after surgery based on pre-treatment nor changes associated with the administration of the isolated Serp-1 protein. Additionally, it was found that Serp-1 improves the histological presentation of acute renal allograft rejection regardless of the status of the microbiome.
Date Created
2022
Agent

Evaluation of the Effects of Corn Flour Consumption on Cardiometabolic Outcomes and the Gut Microbiota in Adults with High Cholesterol

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Description
High fiber diets have been associated with improved cardiometabolic health with specific efforts to lower circulating levels of low-density lipoprotein (LDL cholesterol). Whole grain and grain-based foods are major contributors of dietary fiber in the American diet, of which wheat…

High fiber diets have been associated with improved cardiometabolic health with specific efforts to lower circulating levels of low-density lipoprotein (LDL cholesterol). Whole grain and grain-based foods are major contributors of dietary fiber in the American diet, of which wheat has been extensively studied. Corn, however, has not been well studied for its cholesterol-lowering properties. Further, the mechanisms by which grains improve cardiometabolic health require further exploration with regard to the human microbiome. The objective of this single-blind randomized controlled, crossover trial was to assess the impact of three different corn flours (whole grain, refined, and bran-enhanced refined flour mixture) on serum LDL cholesterol and the gut microbiota diversity and composition. Twenty-three participants were recruited, between the ages of 18-70 with hypercholesterolemia (Male = 10, Female = 13, LDL >120 mg/dL) who were not taking any cholesterol-lowering medications. Participants consumed each flour mixture for 4 weeks prepared as muffins and pita breads. At the beginning and end of each 4-week period serum for cholesterol assessment, anthropometrics, and stool samples were obtained. Serum cholesterol was assessed using a clinical analyzer. Stool samples were processed, and microbial DNA extracted and sequenced based on the 16S rRNA gene. A generalized linear model demonstrated a significant treatment effect (p=0.016) on LDL cholesterol and explained a majority of the variance (R-squared= 0.89). Post hoc tests revealed bran-enhanced refined flour had a significant effect on cholesterol in comparison to whole grain flour (p=0.001). No statistically significant differences were observed for gut microbial community composition (Jaccard and weighted Unifrac) after corn consumption. However, relative abundance analysis (LEfSE) identified Mycobacterium celatum (p=0.048 FDR=0.975) as a potential marker of post-corn consumption with this microbe being differentially less abundant following bran-enhanced flour treatment. These data suggest that corn flour consumption may be beneficial for individuals with hypercholesterolemia but the role of gut microbiota in this relationship requires further exploration, especially given the small sample size. Further research and analysis of a fully powered cohort is needed to more accurately describe the associations and potential mechanisms of corn-derived dietary fiber on circulating LDL cholesterol and the gut microbiota.
Date Created
2022
Agent

Comparing Profiles of the Urinary Microbiome between Patients with Urinary Disease

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Description

Urologic diseases interstitial cystitis (IC), overactive bladder (OAB), and urinary tract infection (UTI) affect tens of millions of people per year in the US alone. The human microbiome consists of a diverse community of bacteria (bacteriome) and viruses (virome) harbored…

Urologic diseases interstitial cystitis (IC), overactive bladder (OAB), and urinary tract infection (UTI) affect tens of millions of people per year in the US alone. The human microbiome consists of a diverse community of bacteria (bacteriome) and viruses (virome) harbored in each individual that contributes to health and disease. Little is known about how the microbiome impacts urinary disorders. Using next-generation metagenomic sequencing, we characterized the urinary bacteriome and virome of patients with urinary disorders (IC, OAB, and UTI) and healthy controls. We show that the bacteriome was distinctly altered in patients by their respective urinary disorder. IC was characterized by a distinct prevalence of the genus Lactobacillus, while OAB was characterized by the genus Bacteroides, and UTI was characterized by Comamonas. IC, OAB, and UTI all also had significantly differed virome profiles from healthy individuals. In particular, we found that Lactobacillus phages were significantly associated with IC and Corynebacterium virus was associated with UTI samples, meanwhile no particular virus was correlated with OAB samples. Overall, we show that changes in the urinary microbiome are associated with incidence and spectrum of urinary diseases. These findings could lead to new microbiome modalities of treatment.

Date Created
2021-05
Agent
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