Undergraduate Journal of Experimental Microbiology and Immunology

Regulators, Processes, and Intricacies of the Innate Immune System with a Focus on the NEMO Complex

2024-06-17T12:04:03-07:00

Insight into immune system regulation is critical to understanding the homeostasis of the immune system, diagnosis of various autoimmune diseases, and cancer prevention measures in the body. The immune system is incredibly complex, from the organ to tissue to cell to molecular level. The innate immune system is contained within each somatic cell and can react to pathogen and damage-associated molecular patterns via pattern recognition receptors. These receptors initiate innate immune pathways that lead to inflammation. Understanding the mechanisms and regulators of the innate immune system and methods by which researchers map innate pathways provides a broader and more holistic context for the immune system's role in many diseases and disorders. To that end, this paper aims to record and analyze multiple proteins and mechanisms of the innate immune system, the role of the leucine rich repeat structure in the innate immune system, structural/functional elements of proteins that contribute to immune regulation of the NEMO (NF-kappa-B essential modulator), standard methodologies for testing signal pathways, and limitations/possible new methods for immune testing.

High Maternal Cortisol Levels During Pregnancy Leads to Dysbiosis in Newborn’s Gut Microbiome

2024-07-03T12:18:11-07:00

Stress during pregnancy can have serious implications for fetal development. Maternal prenatal stress, measured using cortisol concentrations, is a simple measure of stress that is often overlooked when pregnancy outcomes are researched. Heightened stress response through the hypothalamic-pituitary-adrenal (HPA) axis directly affects the maternal gut via glucocorticoid (GR) and mineralocorticoid receptors (MR) on gut epithelial cells. The interaction of cortisol with these intracellular receptors leads to altered epithelial barrier permeability and nutritional availability. Stress results in a shift in maternal gut microbiome composition during pregnancy. This can be transferred to the fetus during birth and prime the newborn’s gut microbiome in a manner that leads to gut dysbiosis during early stages. This review will discuss how the HPA axis is involved with stress responses in the body, the impact of stress on the mother’s gut microbiome, and the transfer of the disrupted mother’s microbiome and/or metabolites to the newborn prior to and during birth.

Chaperone Protein BamB Directly Supports BrkA Translocation and Folding in Escherichia coli

2024-07-12T16:09:19-07:00

Autotransporters are virulence factors expressed on the surface of gram-negative bacteria as part of the type V secretion system. As the name suggests, the surface expression of an autotransporter is facilitated by passage through the protein’s own translocation domain, rather than an additional channel. One such example is BrkA of Bordetella pertussis, a type V autotransporter responsible for adhesion, serum resistance, and cellular invasion. The assistance of chaperone proteins is required to translocate autotransporters across bacterial membranes and enable proper folding into functional structures on the cell surface. However, the exact chaperones required for BrkA surface expression are currently unclear. In this study, a panel of Escherichia coli chaperone knockout strains was transformed with a brkA expression vector to investigate the importance of these chaperones in stable autotransporter surface expression. Through trypsin accessibility assays and western blot analysis we found that BamB may be for the expression of cleaved BrkA on the outer membrane, and may directly interact with the protein during this process. We used cross-linking experiments and found that BamB may directly interact with BrkA autotransporter. This project aims to characterize the functional significance of a wide panel of chaperones with respect to BrkA. Insights derived from this study are relevant to immunotherapeutic and vaccine development, where BrkA is an attractive target due to its surface presentation and its role as a virulence factor.

Escherichia coli Secretion of a Variant in the Bordetella pertussis Autotransporter Protein BrkA Containing a Dual Polyhistidine-Tagged 112 Amino Acid Insert Between Asp-57 And Ala-58 of its Passenger Domain

2024-07-31T16:43:48-07:00

Bordetella pertussis is a Gram-negative bacterium which expresses the Bordetella resistance to killing A (BrkA) protein. BrkA mediates autotransporter functionality and undergoes a processing event to produce a cleaved and uncleaved protein. Despite advancements in the functionality of different regions of this autotransporter, there is a lack of understanding of the mechanistic dynamics driving BrkA translocation. This study aimed to generate a sizable amino acid insert into BrkA passenger domain, analyze the structural layout of the protein, and to determine surface-expression status, regardless of the increase in molecular weight. We hypothesized that inserting a 112 amino acid spacer into the N-terminal domain will not affect BrkA surface expression, but may modulate processing events in an Escherichia coli model. We constructed pSERC, containing two 6x histidine tags within a 336 base pair insert in the brkA passenger region. The amino acid sequence corresponding to the insert revealed passenger residue repetition and potential for a protease cleavage site. Finally, we confirmed that the 112 amino acid insert-containing BrkA passenger retained surface expression in an E. coli model and a processing event. Our findings report that BrkA translocation is retained following a substantial amino acid insertion, and further suggests a potential avenue for studying BrkA surface expression and the complex translocation pathway that the passenger follows from cytoplasm to outer-membrane.

Harnessing the BrkA Autotransporter to Surface Display the Zinc-Binding Domain Tetrahymena thermophila Metallothionein MTT5 on Escherichia coli for Zinc Biosorption Evaluation

2024-07-03T17:07:14-07:00

Heavy metal contamination, particularly zinc, poses significant environmental and health concerns. Surface expression of metal-binding proteins in easily culturable bacteria such as Escherichia coli presents a promising bioremediation strategy to mitigate heavy metal pollution. We investigated the surface expression of Tetrahymena thermophila metallothionein (MTT5), a 10.5 kDa metal-binding protein, using the BrkA autotransporter system in E. coli. We designed a recombinant BrkA-MTT5 plasmid (pADKT5) and confirmed MTT5 expression via western blot analysis. Further investigation utilizing a trypsin accessibility assay suggested the potential for BrkA-mediated export of MTT5. Additionally, the zinc-binding capacity of BrkA-MTT5 was evaluated by an Arabidopsis thaliana root growth assay which demonstrated statistically significant differences between bacteria-treated and non-treated conditions. Overall, these findings highlight the potential for BrkA-mediated export of MTT5 and lay the foundation for further research aimed at harnessing the bacterial surface display system for heavy metal bioremediation.

The DsbA Chaperone Protein May Be Necessary for Outer Membrane Secretion of BrkA

2024-07-27T16:18:43-07:00

Gram-negative bacteria utilize specialized secretion systems to transport proteins across their outer membrane and into the extracellular environment. Among these systems, autotransporter proteins, like BrkA in Bordetella pertussis, autonomously localize to the bacterial cell surface, increasing virulence and inhibiting bactericidal immune responses from hosts. While periplasmic chaperone proteins have been shown to be involved in autotransporter biogenesis, not all their specific roles in BrkA secretion, as it transits through the periplasm, are well understood. DsbA and SurA are two chaperone proteins found in the periplasm of Gram-negative bacteria. SurA has been previously implicated in the secretion of BrkA, while DsbA does not interact directly with BrkA, it’s interactions with SurA and whether these aid in the secretion are unknown. We aimed to explore whether DsbA affects BrkA surface expression in Escherichia coli through loss-of-function experiments. Our results suggest that DsbA may play a role in BrkA secretion. The experimental outcomes indicate a possible role for DsbA in facilitating BrkA secretion, implicating a wider involvement of periplasmic factors in the complex orchestration of BrkA biogenesis and secretion.

Engineering Bordetella pertussis BrkA Autotransporter for Chitin-Binding Domain Surface Expression: Exploring its Potential for Whole-Cell Immobilization

2024-07-03T11:36:52-07:00

The BrkA autotransporter is an important virulence factor in Bordetella pertussis that confers serum resistance and mediates cell adherence. As it is self-secreting and only one protein needs to be manipulated, this makes it a relatively simple secretion pathway that can be exploited to deliver proteins to the surface for applications such as whole-cell immobilization in fermentation bioreactors. In this context, the chitin-binding domain derived from Pseudomonas aeruginosa PAO1 chitinase C emerges as a promising candidate, given its strong affinity for chitin, an abundant polysaccharide, and its pH-sensitive characteristics that simplifies the process of retrieving cells after a bioreaction. As such, we aim to exploit the BrkA autotransporter secretion system to export the chitin-binding domain to the cell surface. By leveraging the binding properties of the chitin-binding domain to chitin, we also aim to explore its potential in whole-cell immobilization. In our study, we successfully engineered a recombinant plasmid (TAAK-A54) by substituting part of the BrkA passenger domain with the chitin-binding domain. A western blot indicated expression of the intracellular chitin-binding domain with a molecular weight of 52 kDa, and the extracellular processed form at 38 kDa. A trypsin accessibility assay confirmed the expression and export of the chitin-binding domain to the cell surface. Lastly, we observed through microscopy that Escherichia coli cells expressing the chitin-binding domain can interact with chitin. Samples had decreased turbidity following incubation with chitin resin, with a maximum 3-fold change relative to the negative control, suggesting that cells were able to be immobilized by chitin. This study enhances the understanding of the repertoire of heterologous proteins that the BrkA autotransporter system can secrete. We demonstrated the feasibility of leveraging the BrkA secretion system for the external presentation of the chitin-binding domain. Our results indicate that the chitin-binding domain is worthy of further study for applications in whole-cell immobilization.

Promoter Mapping Analysis Using a GFP Reporter Plasmid Suggests that the lac Promoter May Drive brka Expression on Plasmid pDO6935 in Escherichia coli

2024-07-09T21:48:45-07:00

Autotransporters (AT) are a class of bacterial proteins that play a crucial role in bacterial virulence and pathogenesis. Understanding these proteins is essential to advance research in infectious disease and identify possible targets for therapeutic interventions. Bordetella pertussis autotransporter, BrkA belongs to the AT-1 subfamily of autotransporter proteins in Gram negative bacteria. Since its discovery, plasmids have been created to study brkA expression in various systems, pDO6935 being one of them. The mechanism by which brkA is expressed in pDO6935 is yet to be fully elucidated. We used a promoter trapping method to map the promoter regions that drive brkA expression. An analysis of pDO6935 gene sequence revealed the presence of a lac operon upstream of brkA. The lac operon is known to exhibit basal expression in the absence of lactose or the presence of a repressor. We studied whether the leaky expression caused by the lac operon may result in the expression of brkA in an Escherichia coli system. In our study, we utilized pSPPH21, a promoterless reporter plasmid that was designed to contain green fluorescent protein (GFP). Using this vector, we inserted regions of putative promoter sequences upstream of brkA in pDO6935. Fluorescence imaging and quantification on a plate reader suggested that the lac operon may be driving brkA expression in pDO6935. We further investigated this argument upon treatment with glucose, a known catabolite repressor of the lac operon. Glucose treatment resulted in repressed levels of GFP, providing additional evidence to our findings.

Mapping the brkA Promoter Region on pDO6935 in Escherichia coli Using pLISA, A Novel Promoterless GFP Reporter Plasmid

2024-06-12T13:19:41-07:00

Bordetella resistance to killing A (brkA) is a serum resistance gene encoding an autotransporter native to Bordetella pertussis, the causative agent of whooping cough (1). Using the expression construct pDO6935, previous studies have demonstrated brkA expression in Escherichia coli. However, the promoter driving brkA expression from this construct is unknown. pDO6935 harbors a lac regulatory region inclusive of a promoter, operator, and catabolite activator protein binding site (CAP BS) upstream of brkA which may be implicated in its expression. The lactose, or lac, operon regulatory elements are known for their ability to control gene expression via transcriptional regulation in E. coli. To characterize the brkA promoter on pDO6935 in E. coli, we created pLISA, a promoterless green fluorescent protein (GFP) vector to be used in a promoter trap experiment. In our work, regions upstream of the brkA locus on pDO6935, either inclusive or exclusive of the lac regulatory region, were cloned into pLISA. GFP expression was observed as an indicator of promoter activity. We showed that GFP expression was induced when amplicons contained the full lac regulatory region. This observation suggests that the lac regulatory region is likely both necessary and sufficient to drive brkA expression from pDO6935 in E. coli - informing future efforts to study virulence gene brkA using E. coli as a model system.

Purified Ag43α Protein as a Potential Method for Preventing Escherichia coli Autoaggregation

2024-06-17T12:06:41-07:00

The phenomenon of Escherichia coli autoaggregation and the subsequent biofilm formation poses significant challenges in both healthcare and environmental spheres, primarily due to enhanced bacterial resistance against antibiotics. Autotransporters, particularly Antigen 43 (Ag43), a prominent outer membrane protein in Gram-negative bacteria like E. coli, facilitate bacterial aggregation through their distinct structural domains. Ag43 consists of a secreted passenger domain (Ag43α), an autochaperone domain and a β-barrel domain anchoring the secreted protein to the outer membrane. Previous studies have established the role of the Ag43α subunit in promoting aggregation through self-recognition. Hence, we hypothesise that the application of exogenously purified Ag43α could interrupt these intercellular interactions, thereby inhibiting autoaggregation. We expressed and purified the Ag43α subunit in BL21(DE3) strain of E. coli cells using the pEEKABOO plasmid from Leong et al. to investigate its influence on the aggregation of the Ag43-expressing DH5α E. coli strain. The results exhibited purified Ag43α protein adopting a stable tertiary structure post-translation. Furthermore, preliminary results from a single-trial aggregation assay revealed that introducing 200 µg/mL of purified Ag43α noticeably attenuated autoaggregation in acidic environments. Our preliminary findings provide a basis for further investigation of the Ag43α protein’s potential in modulating E. coli autoaggregation and a possible avenue for the development of novel anti-biofilm therapeutics.

Chaperone DegP Is Not Necessary for the Polyhistidine Tag-Detected Secretion of Autotransporter BrkA in Escherichia coli BW25113

2024-06-22T14:16:25-07:00

Autotransporters are virulence-associated proteins capable of independent secretion, present on the outer membranes of Gram-negative bacteria. Bordetella pertussis is the causative agent of whooping cough and expresses the 103 kDa type Va autotransporter Bordetella resistance to killing (BrkA), which contributes to the bacterium’s resistance against the bactericidal activity of the complement pathway. The exact folding mechanisms, including the role of periplasmic chaperones involved in the translocation of BrkA, are not well understood. DegP is a highly conserved periplasmic chaperone that has been implicated in the folding and secretion of other autotransporters in Escherichia coli but has yet to be characterized in the expression of BrkA. We therefore hypothesized that DegP would be necessary for BrkA secretion. To investigate this, a 6x histidine tag was inserted in the unstructured region of the BrkA passenger domain for detection by immunoblot analysis. We compared the expression between WT BW25113 and mutant JW0157 (ΔdegP) cells and observed BrkA surface expression in both conditions, indicating that DegP is not necessary for BrkA secretion. This study provides insight into the secretion process of the virulence factor BrkA and bacterial membrane transport systems which holds the potential for the development of new clinical interventions and biotechnological applications.

The lac Promoter May Not Regulate brkA Expression in Plasmid pDO6935 in Escherichia coli

2024-06-26T14:23:57-07:00

BrkA is an autotransporter protein found in Bordetella pertussis that is regulated by the Bordetella virulence gene (Bvg) system. It has been researched in pDO6935-transformed Escherichia coli, with previous research suggesting that the presence of a lac promoter is necessary for BrkA expression in these models. lacI^Q is a constitutively expressed gene found in pHN678 that encodes the LacI repressor, which when in the absence of lactose or analogs, should bind to the operator downstream of the lac promoter, thereby preventing downstream gene expression. Therefore, the objective of this study was to manipulate this mechanism by introducing lacI^Q into a pDO6935-derivative, pENS, to create an IPTG-inducible BrkA expression system in E. coli cells, assuming that the lac promoter was responsible for BrkA expression. We created pTEAM, a construct containing lacI^Q downstream of BrkA on the pDO6935-derived pENS plasmid, which was transformed into UT5600 E. coli cells prior to visualization for BrkA expression. Contrary to our hypothesis, our data showed that BrkA was still being expressed with strong signals by our pTEAM construct in the absence of IPTG whereas our pENS constructs expressed little to no BrkA. This result suggests that the lac promoter may not regulate BrkA expression in pDO6935.

The Distance of the lac Regulatory Region to the Translation Start Codon of gfp on Plasmid pEAH23A Does not Change GFP Expression Levels

2024-06-28T14:02:28-07:00

Bordetella pertussis, the causative agent of whooping cough, uses various strategies to evade host immune defenses, notably through its autotransporter protein Bordetella resistance to killing (BrkA) which blocks complement-mediated killing. Therefore, understanding how brkA expression is regulated is crucial for understanding its role in pathogenesis. Previous research has demonstrated the involvement of the lac regulatory region, located upstream of the brkA locus on the pDO6935 plasmid in brkA expression. When this region was cloned onto a promoterless plasmid, it was able to drive GFP expression. However, the impact of the proximity of regulatory regions on gene expression remains poorly understood. In this study, we investigated how the distance between the lac promoter and a reporter gene, gfp, on a plasmid, impacts GFP expression levels. Utilizing site-directed mutagenesis, plasmids pBRAT24C and pBRAT24D were created, with shorter distances between the lac promoter and gfp relative to pEAH23A. Plasmids were transformed into DH5α Escherichia coli for expression and further analysis. Quantitative GFP analysis showed that pBRAT24C and pBRAT24D induced lower levels of GFP expression than pEAH23A. This suggests that the distance between the lac promoter and gfp does not affect levels of GFP expression and that the sequences found upstream of gfp might affect gene expression through other mechanisms. Further study into the regulatory effects of this region can help maximize BrkA yield from pDO6935, aiding future investigations into its function, structure, and potential as a vaccine candidate.

Mapping Potential Transcription Start Sites of the brka Gene on pDO6935 in Escherichia coli Using the ARF-TSS Method

2024-07-03T12:36:09-07:00

Autotransporters are membrane proteins expressed on the outer membrane in gram-negative bacteria. BrkA is an autotransporter and virulence factor of the whooping cough-causing bacterium Bordetella pertussis. Previous studies aimed at characterizing the autotransporter BrkA have used pDO6935-expressing Escherichia coli cells; however, the promoter driving brkA gene expression in pDO6935-expressing E. coli cells remains unknown. Our study aims to determine the transcription start site (TSS) of the brkA gene in pDO6935 in E. coli (DH5α). In order to map the TSS, we implemented the adaptor- and radioactivity-free identification of the transcription start site method (ARF-TSS) which involves reverse transcribing brkA mRNA into cDNA using a 5'-phosphorylated primer, ligating the cDNA, and PCR-amplifying it before sending it for sequencing to determine the TSS. We were able to more precisely map the region of the plasmid that the transcription start site driving the expression of the brkA gene in pDO6935 in E. coli which is located in a region more than 270 bp upstream of the brkA translation start site. Finding the transcription start site aids in locating the promoter and contributes to our understanding of gene expression and regulation. This knowledge can help improve future studies aimed at characterizing BrkA autotransporter using pDO6935-expressing E. coli cells.

RNA-seq Analysis of Plasmodium falciparum Shows Upregulated Genes Involved in Merozoite Egress and Erythrocyte Invasion

2024-05-14T09:08:36-07:00

Malaria is a febrile disease caused by parasites of the genus Plasmodium. The most severe cases and most of the deaths of malaria are due to Plasmodium falciparum infection. Investigating transcriptional differences in P. falciparum between severe and uncomplicated cases can shed light on malaria pathogenicity and virulence. In this study we used previously acquired publicly available RNA sequencing data from severe and uncomplicated cases of P. falciparum infection. Differential expression analysis of the P. falciparum transcriptome showed that a group of gene products involved in merozoite egress and erythrocyte invasion were significantly upregulated in severe malaria. More specifically, we identified that Subtilisin-like protease 1 (SUB1), merozoite surface proteins 1 and 2 (MSP1 and MSP2), and serine repeat antigen 5 (SERA5) were upregulated in severe malaria. Upregulation of genes in merozoite egress and erythrocyte invasion suggest molecular mechanisms of malaria severity specifically during the blood stages. In addition, upregulation of tyrosine kinase-like protein (TKL) transcription in conjunction with SERA5 regulatory proteins upregulation suggests additional mechanisms of SERA5-mediated malaria severity.

Exploring the Effects of Isotretinoin on the Oral Microbiome: A Comparative Study of Microbial Diversity and Composition

2024-06-18T14:58:14-07:00

Isotretinoin, known by its brand name Accutane, is an oral medication commonly used in the treatment of moderate to severe acne. Previous research has demonstrated the antimicrobial properties of isotretinoin, particularly in modulating cutaneous colonization by the acne linked Cutibacterium acnes, however, its effects on entire microbial communities are poorly understood. Therefore, the objective of this study was to investigate the effects of isotretinoin on the beta diversity of the oral microbiome. The salivary microbiomes of 7 individuals who have previously taken isotretinoin but are no longer taking it were compared to 27 control individuals with no history of isotretinoin usage. 16S sequencing followed by non-metric multidimensional scaling showed no significant difference in oral microbiome beta diversity between individuals who have taken isotretinoin and those who have not. Differential abundance analysis revealed past isotretinoin users exhibited significantly lower abundances of Actinobacteria taxa including Actinomycetales and Micrococcaceae related species. While bacterial alpha diversity was similar between study groups, oral community richness within the Actinobacteria phylum was significantly lower among isotretinoin users. These findings provide further insight into the long-term effects of isotretinoin on the human body and characterize its impact on the oral microbiome for the first time. However, additional research is needed to explore the underlying mechanisms by which isotretinoin influences microbial diversity as well as the broader implications of such alterations in microbiome composition.

Sex Influences Gut Microbial Composition in Mice with Familial Dysautonomia but is not the Primary Determinant of Microbial Functional Diversity

2024-08-16T17:05:05-07:00

Familial dysautonomia is a genetic neurological disease characterized by impaired nervous system functions due to a mutation of Elongator acetyltransferase Complex subunit 1. Previous studies have demonstrated that the severity of familial dysautonomia is associated with variations in the gut microbiota and metabolite profiles in both human patients and mice. However, the mouse model for familial dysautonomia has yet to be characterized in terms of demographic factors (i.e. sex and age) associated with their gut microbial dysbiosis and metabolic composition. Hence, we aim to investigate the variables associated with gut microbial dysbiosis and how they impact gut microbial dynamics and metabolic functions in mice with familial dysautonomia. First, through univariate regression analysis with Bray-Curtis distance and principle coordinate analysis, we determined that sex was the strongest indicator correlated with compositional variations. Next, our taxonomic composition analysis revealed a decrease in the abundance of Bacteroidota and increased Firmicutes in male familial dysautonomia mice compared to females. Furthermore, indicator species analysis identified signature genera of class Clostridia, that were representative in male familial dysautonomia mice. Despite no significant sex differences across metabolic pathways from a predictive functional analysis, correlation analysis identified a strong association between genera Romboutsia and creatinine degradation II pathways that were shown to be upregulated in males compared to female FD mice. Ultimately, our findings presented that microbial dysbiosis is intricately linked with sex in the familial dysautonomia mouse model. Further characterization of the mouse model allowed insight into its representation of FD and applicability to human patients.

Dopaminergic Therapeutics for Treating Parkinson’s Disease Were Associated with a Shift in the Gut Microbiota to Resemble Healthy Individuals

2024-07-12T08:17:43-07:00

Parkinson’s Disease (PD) is a neurodegenerative disorder associated with dopaminergic neuron loss, leading to dopamine dysregulation. Dopaminergic therapeutics are often administered to restore dopamine levels and have been associated with changes to the gut microbiota. Through a secondary data analysis of a cross-sectional cohort of PD patients, we aimed to investigate changes in the gut microbiome associated with the use of four dopaminergic drugs (entacapone, pramipexole, rasagiline, amantadine). Although the use of dopaminergic therapeutics was not associated with compositional alterations to the microbial diversity of PD patients, we observed changes to specific taxa. Amantadine and pramipexole therapeutics were both associated with a core microbiome that contains Faecalibacterium – a genus contained in the core microbiome of healthy individuals but absent in untreated PD patients. Furthermore, entacapone and amantadine use was associated with taxa that are indicative of a healthy gut microbiome, including Lachnospiraceae and Colidextribacter. We also identified three genera that were differentially abundant with dopaminergic drug use. Dopaminergic therapeutic use was generally associated with increased Bifidobacterium, decreased Prevotella, and increased Akkermansia. While increased Bifidobacterium is associated with a healthier gut microbiome and Akkermansia is associated with gut dysbiosis, the effects of Prevotella remain unclear. Our findings suggest that dopaminergic therapeutics are associated with alterations in the gut microbiome of PD patients that provide an overall benefit to the host. Future studies could incorporate higher resolution analysis at the species level and explore causational effects of dopaminergic drugs in a prospective study.

Eczema May Be Masking the Effects of Multiple Sclerosis on the Gut Microbiome

2024-07-28T23:03:45-07:00

Multiple sclerosis is a chronic autoimmune-mediated neurodegenerative disease with no cure leading to physical disability and cognitive impairment. Eczema patients often have an altered gut microbiota and increased susceptibility to autoimmune illnesses such as multiple sclerosis. In addition, these patients with multiple sclerosis and eczema are generally burdened with taking a higher number of medications, which can impact gut microbial diversity. However, the impact of eczema on multiple sclerosis and corresponding medication intake in treatment has yet to be fully established. Here, we investigate gut microbiome composition in multiple sclerosis patients and healthy controls with and without eczema that take varying medication types. QIIME2 was utilised to process 16S rRNA gut microbiome sequences from the International Multiple Sclerosis Microbiome Study prior to analyses on alpha and beta diversity, core microbiome, and indicator species with R. Interestingly, significant differences in beta diversity between multiple sclerosis patients and healthy controls were only observed if patients did not have eczema. Similarly, significant beta diversity differences between patients taking or not taking over-the-counter medication were only observed if patients did not have eczema. Eczema was concluded to potentially mask the effects of multiple sclerosis and over-the-counter medication use, creating a mediating effect on the gut microbiome. Indicator taxa analysis of multiple sclerosis patients reveals 28 indicator species for eczema, suggesting a unique gut microbial community in eczema patients. This suggests that additional consideration should be taken when deciding treatment courses for multiple sclerosis patients with and without eczema.

The Diet of the Hadza Tribe is Higher in Gut Microbial Diversity, but Lower in Functional Diversity when Compared to a Westernized Diet

2024-07-03T11:49:06-07:00

The Western diet has recently been a point of scrutiny in the scientific community due to its association with various metabolic diseases such as obesity, type 2 diabetes, cardiovascular disease, and certain cancers. However, the unique hunter-gatherer diet of the Hadza tribe has been noted to be protective against such diseases, which has been partially attributed to their unique gut microbiome. While previous studies have compared the gut microbiomes of the Hadza to those of Western countries, very few have looked into its relationship to the gut microbiome’s metabolic pathways and functional diversity. In our study, we confirm the literature’s finding that the Hadza microbiome, when compared to a westernized gut microbiome, is higher in alpha diversity and demonstrates high beta diversity. Additionally, we found multiple beneficial taxa to be present in higher amounts in the Hadza gut microbiome with more dysbiotic species being present in the westernized one. Further, our study indicates that the Hadza surprisingly demonstrated lower functional diversity, while the westernized dataset contained more upregulated metabolic pathways.

Nutrients May Play a Complex Role in Influencing Microbiota Composition and Function in Parkinson’s Disease Patients

2024-07-11T11:27:47-07:00

Parkinson's disease is a neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons, leading to symptoms such as tremors and gastrointestinal dysfunction. Previous studies have shown reduced gut microbial diversity and imbalances in Parkinson’s disease patients including increases in inflammatory microbes and decreases in commensals. However, the impact of dietary macronutrients and micronutrients on gut microbiota composition in Parkinson’s disease patients remains unclear. By analyzing published microbiota data containing nutrient intakes of Parkinson’s disease patients and non-Parkinson’s disease controls, we aimed to identify nutrients that are associated with gut microbiota composition in Parkinson’s disease patients. We observed correlations between certain nutrients and changes in beta diversity among Parkinson’s disease patients. We then segregated the Parkinson’s disease and control data based on consumption of each of these identified nutrients and performed taxonomic analyses and functional pathways analysis. These analyses compared the upper and lower nutrient consumption groups for Parkinson’s disease and control populations. The core microbiome and indicator species analyses revealed that microbial composition is more responsive to nutrient intake in the control group than in Parkinson’s disease patients, with a greater number of core ASVs and indicator taxa identified in control groups. Differential abundance analysis further showed that high nutrient intake generally led to the upregulation of more ASVs, with some exceptions, such as coffee in PD patients and NSP in non-PD controls, where downregulation was more prominent. The functional pathway analysis identified that nutrients have a greater metabolic effect on Parkinson’s disease patients than control. Ultimately, our survey into how nutrients drive changes in microbial diversity in Parkinson’s disease patients may help guide future research into nutrient intake catered to the prevention and management of Parkinson’s disease.

Bacterial Composition and Metabolic Pathways Differ in Six-Month-Old Infants from Peru and California, USA

2024-07-22T14:59:19-07:00

The infant gut microbiome rapidly develops with age and is influenced by various factors. The potential influence of an infant's geographical origin on these adaptations should be considered. We compared six-month-old infants from two distinct locations — Iquitos, Peru, and San Diego, California, USA —to identify geography-related differences in infant gut microbiomes. This comparison was conducted by analyzing microbial diversity, composition, and functional phenotype in fecal samples. Beta diversity analysis suggested statistically significant differences in microbial communities between the two infant cohorts. Functional analysis using the PICRUSt2 software revealed an overrepresentation of different bacterial metabolic pathways associated with infant development in both infants from Peru and California. However, taxonomic results showed that the bacteria commonly associated with these metabolic pathways, namely, Faecalibacterium, Lactobacillus, and Bifidobacterium, were more abundant in the infants from Peru than from California. These findings varied in similar patterns between the two cohorts, suggesting a potential difference in the microbiome of the infants from these geographical locations. This highlights the need for further studies to directly uncover and characterize microbial variations between infants from different regions through direct location comparisons between multiple groups as well as data collection in understudied geographical locations and demographics. Ultimately, this may improve the quality of results and encourage further studies on diverse geographical locations and demographics to expand our knowledge on the infant intestinal microbiota and different functional activity, worldwide.

Ironing Out Anemia: High Inflammation Modulates Microbial Composition and Indicates the Activation of Cellular Stress Responses in Infected Anemic Infants

2024-07-12T15:48:00-07:00

Iron Deficiency Anemia (IDA) is a significant global health challenge impacting cognitive, physical, and behavioral development, particularly in infants. Despite the current knowledge on anemia and its manifestations, the role of the gut microbiome and its relationship with systemic inflammation and anemia have yet to be explored. This study explores the relationship between anemia, systemic inflammation, and the gut microbiome in infants using a dataset from McClorry et al. Our analysis revealed that neither infection status nor inflammation levels significantly altered microbial diversity, however, differentially abundant genera were related to inflammation levels in infected anemic infants. Metabolic pathways analysis further identified the upregulation of pathways involved in energy production and immune response, emphasizing the systemic effects of inflammation in anemic infants. Our findings underscore the potential of microbial interventions in managing inflammation-related anemia, highlighting the need for further research into the microbiome’s role in inflammatory processes.

Smoking in the Context of Westernized Diets is Associated with a More Volatile Microbiome and Predicted Gut Function

2024-07-11T12:16:26-07:00

With the growing consumption of a high-fat Westernized diet in Latin America, a similarly Westernized microbiome is becoming predominant. Such a microbiome brings associations of dysbiosis and harmful metabolic products, as well as the risk of cardiometabolic disease – symptoms also associated with cigarette smokers. It is currently unclear whether the combination of a Westernized diet and cigarette smoking results in a compounded effect on the gut microbiome. In this study, we aim to investigate the synergistic effect of smoking with high blood levels of low-density lipoprotein (LDL) on the gut microbiota composition and function. Our analysis was performed to explore this potential interaction between two causes of health concern and their implications for microbiome-influenced health. Using data from a 2018 microbiome study by de la Cuesta-Zulugala et al., our analyses show no significant changes in alpha diversity metrics between smoking & non-smoking individuals and their blood LDL levels. However, at both a taxonomic and functional level smokers’ microbiota appear significantly more affected by blood LDL than those of non-smokers, which were comparatively stable. Changes of note include significant increases in hallmark taxa of Westernization (Bacteroides and Clostridia), as well as downregulation of pathways relating to degradation of both aromatic compounds and D-glucarate. We postulate that the former may be in response to polycyclic aromatic hydrocarbons found in cigarette smoke, while the latter is implicated in regulation of blood cholesterol levels, suggesting a more complex interplay between smoking and measured blood LDL. Together, our findings suggest that smokers experience a more volatile gut microbiome that can be mediated through informed dietary choices.

Parkinson’s Disease Status but not High Sodium Consumption is Correlated with Alterations in the Gut Microbiome

2024-08-02T11:24:12-07:00

Globally, Parkinson's disease (PD) is emerging as the most rapidly growing neurological disorder. PD is associated with a strongly altered gut microbial composition, which may promote disease in part by increasing inflammation. A high sodium diet (HSD) has been shown to alter gut microbial composition in a pro-inflammatory manner, and thus our study aimed to determine whether people with PD who had HSDs experienced more dysbiosis than those with lower sodium intake. If a correlation between HSDs and an exacerbated PD gut phenotype exists, adopting low-sodium diets could emerge as a viable strategy to mitigate PD gut dysbiosis and alleviate associated downstream symptoms. We examined differences in gut microbial diversity and composition related to an HSD within a cross-sectional cohort of 281 individuals both with (n=182) and without PD (n=99). Our approach involved employing alpha and beta diversity analyses alongside differential abundance analyses at the amplicon sequence variant (ASV) level. An HSD was associated with significant differences in the gut microbial composition within the control subjects, but not those with PD. Thus, our findings suggest that the factors underlying the distinct gut microbial profile associated with PD appear to exert a more pronounced influence than the impact of sodium. Based on our findings, there is no compelling evidence to advise individuals with PD against consuming HSD for alleviating gastrointestinal dysbiosis. Overall, our research provides more insight into the correlation between sodium and the gut microbial composition of individuals both with and without PD and establishes novel avenues for future research.

Low Serum Saturated Fatty Acid Levels Positively Associate with Microbiota Diversity and Metabolic Pathways in Parkinson's Disease Patients

2024-07-12T16:15:28-07:00

Parkinson’s disease is the fastest-growing neurodegenerative disease worldwide. Gut microbiome dysbiosis can precede the onset of Parkinson’s disease symptoms by 20 years. The ketogenic diet has shown beneficial impacts as an intervention in the treatment and modulation of the microbiome in Parkinson’s disease. While the ketogenic diet improves Parkinson’s disease symptoms, it increases the serum levels of saturated, monounsaturated, and polyunsaturated fatty acids. The current literature indicates conflicting results with the increase in intake and serum levels of fatty acids and the gut microbiome in Parkinson’s disease. We analyzed the data of 197 Parkinson’s disease patients and 103 healthy controls to unveil associations between serum levels of saturated, mono-unsaturated, and polyunsaturated fatty acids and the microbiome. Our results indicate that saturated fatty acids have a weak but statistically significant positive relationship with the Shannon diversity of the gut microbiome in Parkinson’s disease subjects. Mono-unsaturated and poly-unsaturated fatty acids were not significantly associated with the microbiome diversity. Additionally, we identified low saturated fatty acids associated with the Akkermansia, Bifidobacterium, Faecalibacterium, and Haemophilus genera, with implications in Parkinson’s disease progression and gut dysbiosis. Our analysis also shows low saturated fatty acid positively associates with metabolic pathways such as menaquinol and L-methionine, both having been highlighted as beneficial for Parkinson’s disease. Together, our study indicates that low levels of serum saturated fatty acids are associated with specific genus and pathway changes known to have a positive effect on individuals with Parkinson’s disease.

Individuals in Shared Dormitories who Rarely Wash their Sheets are Associated with Greater Sex-Specific Microbial Changes Compared to Individuals who Wash their Sheets Frequently

2024-06-25T08:37:20-07:00

The skin is the largest organ in the human body whose surface contains a diverse microbial community that contributes to human health by harboring and protecting against pathogens. The composition of the microbial community on the human hand is unique as it has frequent and direct interactions with the surrounding environment. Various factors impact the microbial species present on the dominant hand of an individual, with a primary intrinsic factor being sex and an extrinsic one being hygiene practice. While hygiene practices like hand washing have been shown to decrease overall microbial load, there is a current knowledge gap on how other hygiene practices, such as sheet washing, intersect with sex to impact the skin microbiome. This study therefore explored the impact of sex and the hygiene factor of sheet washing frequency on the microbial composition of hands among individuals residing in shared dormitories. Through microbial diversity and abundance analyses, our findings suggest that sex is a greater driver of hand microbial composition than sheet washing frequency, but that sheet washing frequency still has an effect, as less frequent sheet washing is associated with greater variations in hand microbial composition. Overall, the findings from our study contribute to the growing field of research on how hygiene habits influence the human microbiome in a sex-specific manner, providing a platform for further investigations on the effects of these intersecting factors on health outcomes.

Dogs Are a Poor Taxonomic Model for Human Inflammatory Bowel Disease, but Are Potentially Functionally Relevant for Human Ulcerative Colitis

2024-07-22T14:49:22-07:00

Inflammatory bowel disease (IBD) is a chronic and relapsing inflammatory condition affecting the gastrointestinal tract. Human IBD consists of two major subtypes, Crohn’s Disease (CD) and Ulcerative Colitis (UC) which continue to have large global health and economic implications. Although dogs have previously been investigated as a potential research model for human IBD, their utility for specific subtypes has not been adequately considered. In this study, we explored the microbial diversity, abundance, and functional pathways of the gut microbiota in IBD dogs against UC and CD humans, to investigate if IBD dogs may serve as a research model for a particular human IBD subtype. Comparative analysis of combined 16S ribosomal RNA data from IBD dogs and humans revealed a difference in gut microbial composition between dogs and humans, further supported by a lack of common taxonomic groups in both core microbiome and indicator species analyses. Despite significant species-driven compositional differences, predictive functional pathway analysis displayed similarities in shared metabolic pathways of IBD dogs and UC humans. These findings suggest that while dogs may not serve as relatable taxonomic models for human IBD subtypes, they may hold functional relevance for human ulcerative colitis.

Soil Microbiome Beta, Taxonomic and Functional Diversity in British Columbian Logged Douglas-Fir Stands Differs Across the A and O Horizon

2024-08-02T10:57:19-07:00

Soil bacterial composition varies based on a wide variety of factors and is responsible for much of soil nutrient cycling. To date, the correlations between microbial community composition, functional potential and different soil layers or horizons are not well studied. We investigated the top two organic soil layers, the A horizon and O horizon, in sites of logged British-Columbian Douglas-fir trees and their correlations with microbial diversity. We investigated microbial diversity through alpha diversity, beta diversity and core microbiome analyses. Additionally, we aimed to characterize the microbiota through indicator taxa analysis and PICRUSt2 analysis to determine specific taxonomic and functional differences between the horizons. We found no significant differences in alpha diversity across the A and O horizons whereas in our beta diversity analyses we found clustering within each respective horizon. Furthermore, core microbiome investigations revealed some overlap at the genus level along with greater taxonomic diversity in the O horizon. We also found differences in the top indicator taxa for each horizon using indicator taxa analyses. This was further supported by functional pathway analysis. 407 MetaCyc functional pathways were significantly differentially abundant between horizons, indicating distinct functional niches. These results affirm the clear differences between the microbial diversity and community composition in the A and O horizons, indicating the need for more microbial diversity research regarding horizon depth and logged soil. This will hence improve our understanding of microbial community formation and the potential impact of environmental and human-made factors.

Surgical Intervention Correlates with Reduced Bacterial Diversity in the Gut Microbiome of Crohn's Disease Patients Exhibiting Low Levels of Inflammation

2024-07-22T11:34:04-07:00

Crohn’s Disease is a type of inflammatory bowel disease characterized by dysbiosis and chronic inflammation of the gastrointestinal tract. First line therapies utilize anti-inflammatory treatments alongside endoscopies to manage and monitor disease course. Biomarkers such as fecal calprotectin are being investigated to measure inflammation and predict disease recurrence, and offer a less invasive method to track disease progression. Surgical resection serves as a last resort treatment to medical therapies, but this alternative has been shown to alter microbial diversity post-operatively. This study aims to identify the link between inflammation and microbial composition in Crohn’s Disease patients with or without surgical resection. Diversity analysis revealed reduced alpha diversity in Crohn's Disease patients following surgical intervention. This observation was minimal in patients with pre-existing inflammation compared to those without inflammation. In parallel, we ran the same diversity analysis based on disease severity. We found that inflammation masked the reduction in alpha diversity in patients with more severe disease, consistent with the effects of inflammation on surgical resection. Additionally, indicator species analysis revealed reduced abundance of anti-inflammatory taxa in patients following surgical resection. These findings provide insight into the post-operative intestinal environment, and can help inform post-operative care and limit significant alterations to the gut microbiome in an already dysbiotic environment.