Current GEM Studies

Did you know that the first person to have their genome sequenced was a volunteer from Buffalo, NY?

Genes for Buffalo is a research project designed to continue this proud tradition by collecting saliva samples from adult Buffalonians (18 years or older) to contribute to the worldwide search for causes and cures of human diseases.

By creating a database of DNA sequences (genetic material) from Buffalonians and anonymously link it to a list of diseases they suffer from, we will be able to search for sequences that affect many medical conditions. The database is made such that no personal information like your name or address or doctor is kept, and so that your personal medical information is not viewable. This helps ensure your privacy.

Currently, this research study is being carried out at the UBMD Neurology Clinic at the Conventus Medical Building, with plans to expand collection points soon! If you have any questions or want to learn more please contact the GEM director of outreach activities at (716) 881-7510 or bebrace@buffalo.edu. For any further inquiries, please contact the Principal Investigator for the study, Richard Gronostajski at (716) 829-3471 or rgron@buffalo.edu. 

ABSTRACT

It is established that inflammation in the central nervous system increases susceptibility to the onset of seizures through a variety of mechanisms. Peripheral inflammation can also cause seizures and even though the underlying mechanisms are not as well understood, these data indicate that communication between the two compartments is constant and an important determinant of neurological function and health. Given the increasingly recognized role that the intestinal microbiota plays in shaping a host’s inflammatory environment, we hypothesized that changes in the composition of the microbiota would impact the onset, strength, and/or duration of seizures. Using a well-established model of inducing seizures by reducing inhibitory neurotransmission, we found that mice treated with a cocktail of antibiotics that depleted their intestinal microbiota developed seizures that were less severe and took longer to initiate. These data are significant because they reveal that microbes in the gut can modulate synaptic activity and seizures. These data also suggest that the gut microbiota may be a therapeutic target for the treatment of seizures and epilepsy. Here, we will address the following two questions. First, what type(s) of organisms in the gut microbiota affect(s) seizures and do they affect seizures induced by multiples types of stimuli? Second, are the structures of synapses affected in mice when their microbiota is depleted? Successful execution of these experiments will generate preliminary data necessary for a competitive NIH R01 grant proposal.

ABSTRACT

The omnibus hypothesis of our research is that oral and gut bacteria are associated with the onset and progression of atherosclerotic cardiovascular disease (CVD), as evidenced by microbes from these sites being present in the atheromas. Our long-term goal is to understand how biofilms may contribute to atheroma formation and instability as a basis for targeted interventions to reduce the effects of the biofilm on CVD. Potential mechanisms by which bacteria promote atherosclerosis include foam cell formation, inhibition of HDL reverse cholesterol transport, and induction of proinflammatory signaling. Potential mechanisms precipitating atherosclerotic clinic events, such as myocardial infarction, include biofilm dispersion within an existing atheroma, triggered by iron-dependent increases in norepinephrine, degradation of the fibrous cap, platelet aggregation and thrombogenesis.

Previous work, including ours, using older bacterial DNA technology indicate oral and other bacteria exist in atheroma. We propose to use next generation sequencing to characterize the composition of the microbiome in human atheroma; to compare this with the microbiome in oral, gut and skin samples from the same individuals; and to determine the source of the organisms in the atheroma based on DNA sequence similarity using oligotyping. Additionally, we will identify and localize bacteria in atheroma by the fluorescence in situ hybridization (FISH) methodology to determine the extent they can be characterized as biofilms. To further understand how the atheroma microbiome relates with CVD, we will explore relationships between microbiome characteristics and environmental factors including smoking, serum lipids, and periodontitis. This pilot study will inform sample size and power requirements for a larger grant submission to NHLBI.

ABSTRACT

Trypanosoma brucei, the causative agent of Human African Trypanosomiasis (HAT), encounters dramatically different environments as it traverses its life cycle. For example, the human bloodstream provides 37oC and high glucose, whereas the 27oC mid-gut of the tsetse fly insect vector is depleted of glucose but rich in proline. The parasite responds to these environmental changes by radically altering its physiology and morphology. Remarkably, RNA polymerase II-mediated transcription is constitutive and unregulated in T.

brucei. Consequently, the continuous and profound metabolic and morphological switches that take place during the life cycle are regulated not by transcription factors, but by RNA binding proteins (RBPs) that modulate mRNA stability and translational efficiency. We discovered DRBD18, an RBP that is essential for survival of both bloodstream form (BF) and insect vector procyclic form (PF) T. brucei. DRBD18 knockdown leads to significant changes in the levels of ~1000 mRNAs in PF; however, its impact on the transcriptome of the disease-causing BF is unknown. Many mRNAs that are most significantly affected by DRBD18 knockdown in PF encode RBPs and protein kinases, likely positioning DRBD18 at the apex of numerous regulatory cascades. Additionally, transcriptome studies in PF implicate DRBD18 in promoting the PF phenotype, indicating that this key RPB must impact distinct pathways in PF vs. BF, presumably due to environment- specific protein-protein interactions and/or posttranslational modifications. Here, we will test the hypothesis that DRBD18 differentially regulates the T. brucei transcriptome in BF and PF life cycle environments by direct association with distinct target mRNAs and cis-acting elements.

ABSTRACT

A hypervirulent pathotype of Klebsiella pneumoniae (hvKP) is undergoing global dissemination from the Asian Pacific Rim (1, 2). In contrast to the usual healthcare-associated venue for “classical” K. pneumoniae (cKP) infections in the West, hvKP causes serious, life and organ threatening infections in younger, healthy individuals from the community and has the ability to metastatically spread from sites of infection (1, 3-5). Further, hvKP are becoming increasingly antimicrobial resistant including acquisition of carbapenemases (6, 7). The prospect of a hypervirulent pathogen that is capable of causing severe infection in healthy, ambulatory individuals is alarming; the widespread evolutionary confluence of the hypervirulence of hvKP with the drug resistance of cKP is frightening. Colonization is the first step requisite for infection (8, 9), but a knowledge gap exists on the prevalence of this event. Although all ethnic groups have suffered infection due to hvKP, individuals of Asian extraction appear to be disproportionately affected. Presently it is unknown whether this is due to more frequent geospecific exposure, an underlying genetic abnormality, the development of autoantibodies directed against an unknown host defense factor, high-density colonization, variable colonization resistance, or another factor. Establishing colonization prevalence and density at various geographic locales will assist in resolving these hypotheses and guide future direction. We propose a novel approach to efficiently generate these critical data using a bioinformatics approach. These data will have inherent value by lending insights into the risk of hvKP infection, assisting in understanding its pathogenesis, identifying populations at risk, and generating insights on prevention. Further, these areas are fertile and critical topics for future study. Data generated from proposal will also serve as the foundation for successful external funding.

Abstract

Pathogenic fungi pose a serious danger to immunocompromised people. To establish an infection, these microbes interpret and respond to various nutrient environments in the host by establishing appropriate gene expression patterns. One way of sensing nutrient availability is through the levels of common metabolites, which also serve as cofactors for enzymes that modulate genome architecture and gene expression. For example, the metabolite NAD+ is required by enzymes called sirtuins, which generate a compact chromatin state that represses gene expression. However, it remains poorly understood how metabolites such as NAD+ act as signaling molecules. To elucidate this pathway, the goals of this project are to determine (i) which genes are regulated by sirtuins and hence are activated when intracellular NAD+ is low, and (ii) how nutrient availability in the environment influences intracellular NAD+ concentrations. For these studies, we will focus on three pathogenic fungi that cause serious morbidity and mortality among immunocompromised people. These microbes must adapt to a variety of nutrient environments within a human host, and it is likely that sirtuins contribute to this process. These studies will advance our understanding of how NAD+ acts as an intracellular signaling molecule that is sensed and interpreted by sirtuins to enable cells to respond appropriately to their nutritional environment. In addition, this work will provide new insights into how pathogenic fungi colonize their human hosts.  This collaborative project draws on our expertise in sirtuins (Rusche), fungal pathogenesis (Panepinto), and computational analysis of sequencing data (Liu).

ABSTRACT 

Autoimmune and inflammatory disorders, including psoriasis and eczema, affect hundreds of millions of people. However, there are significant gaps in knowledge regarding the genetic and environmental causes underlying the susceptibility to these diseases. From an evolutionary perspective, skin harbors some of the most significant adaptations due to the wide range of environments that humans colonized. Our preliminary data showed that genetic variants leading to these adaptations are associated both with autoimmune/inflammatory skin diseases and with microbial composition of the skin. Therefore, our central hypothesis is that adaptive human genetic variations involved in skin function are associated with susceptibility to autoimmune/inflammatory disorders through their influence on the skin microbiome by indirectly inducing specific immune responses. To test this hypothesis, we will undertake a comprehensive strategy to analyze a) host genetic variation, b) transcriptomes, and c) microbiomes from non-affected and from affected skin samples from multiple psoriasis and eczema patients. Our proposal is innovative at three levels. First, by combining evolutionary and biomedical perspectives, it will address why skin autoimmune/inflammatory disorders are common and not eliminated from the population by natural selection. Second, it will incorporate underappreciated genomic structural variants, which are major contributors to disease susceptibility. Third, our sampling strategy and utilization of cutting edge -omics tools, will allow measuring the impact of host genetic variation to “healthy” and “affected” microbiomes in two common skin diseases. This innovative collaboration between two leading UB laboratories will lead to at least one major publication, and preliminary data for an R01 grant.

Abstract:

Our recent study revealed a severely reduced number of gut species in obese patients. This internal ecological disaster could contribute to the development and maintenance of obesity and its comorbidities. Abundant representation of conventional probiotic species in the gut of obese patients explains the ineffectiveness of these species in the treatment of obesity and calls for novel probiotic species for obese patients. Keystone species are those species whose activities and abundances are required for the integrity and stability of the community. Our preliminary data demonstrated extensive interactions among gut species, which predicts that several lost/decreased species are keystone species in the gut. Therefore, we hypothesize that probiotic treatment using the reduced/lost keystone species will return the altered microbiome toward normal, and consequently help to reduce weight and to prevent/reverse the mass extinction of species in the gut of obese patients. Our Specific Aims are: Aim1, to determine the dynamic compositional change of the gut microbiome during the development of obesity in rats fed a high fat diet; Aim 2, to identify the keystone species using both the correlation method and the method of interaction analysis (Lotka-Volterra model). Accomplishing the proposed studies will establish a method for the identification of the keystone species in the gut microbiome using cross-sectional data. We will then evaluate the therapeutic effect of the keystone species in the treatment of obesity and its comorbidities in animal models including a gnotobiotic mouse model carrying a humanized microbiome.

Abstract:

There are several lines of evidence to suggest that the gut microbiota is likely altered in patients with chronic kidney disease (CKD), and it has been well known that there is an increased risk of worsening renal function and CKD progression in patients on proton pump inhibitors (PPIs). Several studies have found that PPIs affect gut microbiome through different mechanisms including altering stomach acidity affecting its ability to defend against ingested microbes, increasing colon transit time and direct effects on bacterial survival all through its effects on both bacterial and colonic H+K+ ATPase. This PPI related gut microbiota dysbiosis have been linked to increased risk of autoimmune diseases and colitis. We have recently shown that NHANES participants who are infrequent yogurt/probiotics consumers are more prone to proteinuric kidney diseases compared to frequent users. We propose in this project to perform a cross sectional analysis of gut microbiota in diabetic nephropathy (DN) patients consuming PPIs and compare it to nonuser DN controls.

Abstract:

We propose to carry out a prospective, longitudinal pilot study to analyze the pattern of the human pregnancy ­related (cervical, vaginal, blood, placental and cord), oral, and gut microbiota and their association with pregnancy outcomes.  Current knowledge of the human microbiome and its relationship to health and disease is rapidly increasing¹, and the relationship of pregnancy-related microbiome to pregnancy outcomes, especially preterm birth, is an area under intense study right now.  Recent studies have shown that the placenta and placental membranes, once thought to be part of a sterile environment, may be subject to bacterial invasion on a regular basis. Whether this invasion is pathologic or beneficial, or both, is unknown and part of the impetus for this study.  We hypothesize that colonization/invasion of the human placenta by microorganisms is common, and the frequency, origin and species of this invasion may be associated with pregnancy outcomes.  This study will entail recruitment of pregnant women in the first trimester of pregnancy, assessment of medical and oral health and collection of biologic samples for microbial assessment at 3 points during prenatal care and at the time of delivery.  Next we propose to identify the microbiota of the above samples through 16S RNA meta-genomic sequencing; collection of data on pregnancy outcomes, including gestational age at delivery, birth weight, complications, and delivery mode; and correlation of outcomes with microbial patterns.  We also will correlate the oral disease measurements and pregnancy outcomes with the microbiological results.  Finally, we will compare the placental flora with that from other sites in the population to determine which is (are) the likely sources for the placental flora.

Abstract:

Of all human-associated microbiomes, the human oral microbiome is comprised of one of the most diverse sets of organisms. In addition to bacteria, the human oral microbiome contains viruses, fungi, protozoa and archaea. Although it is well known that certain microbiome bacteria are responsible for the two commonest bacterial diseases of man, dental caries (tooth decay) and the periodontal (gum) diseases. While these diseases are clearly caused by bacteria, the onset and severity of dental disease is not yet predictable from the composition of the oral bacteriome. This observation suggests that complex interactions between the disease-causing bacteria and other microbiome components may regulate the population and disease-causing capacity of these bacteria. Protists can significantly impact a microbiome’s bacterial population size and/or composition. However, studies of the oral microbiome to date have been severely biased towards characterization of the bacterial components, and to a lesser extent, fungal species. Therefore, given the potential importance of bacterial-protist interspecies interactions to oral microbiome functions, we propose to identify and quantify the protist population in the human oral microbiome and examine its potential role in regulating the oral bacteriome. We assert that understanding the ecology of bacterial-protist interactions inherent within the oral microbiome will provide formidable insights that will aid in diagnosis of oral diseases, and point to interventions that may increase disease treatment and/or prevent disease development.

Abstract: Individuals with COPD have a higher risk for lung cancer that is not commiserate with their smoking history or age. Our hypothesis is the increased risk of Lung cancer with those with COPD is possibly due to interaction of the airway microbiome and the host immune-inflammatory response, which enhances the malignant conversion of airway epithelium. We will test this hypothesis utilizing sputum samples obtained during a VA funded study. In this prospective longitudinal study of bacterial infection in COPD from 1994-2014, in which, clinical information, sputum and serum samples were collected monthly from participants, 13 of 182 participants, were diagnosed with lung cancer during the study. We will match these 13 individuals with those who did not develop lung cancer, but were similar in age, smoking habits, lung function and duration of follow-up. We propose to analyze the sputum microbiome and epigenetic biomarkers to identify determinants that are associated with the development of lung cancer.

In the proposed project, we would study the microbiome in these sputum samples with16S sequencing, and assess host epigenetics by examining the DNA methylation profile with next generation sequencing capabilities at the UB Genomics and Bioinformatics Facility. We will identify microbiome and methylation interactions that are associated with development of lung cancer. We have already investigated and published the sputum microbiome in a subset of the stored sputum samples obtained immediately before and after COPD exacerbation from this sample collection, demonstrating the suitability of the stored samples for DNA extraction for microbiome and genetic analysis.