Volker Mai, PhD, MPH – Grants and Contracts
Advanced Computational Algorithms for Deep Interrogation of Microbial Communities Using Millions of 16S rRNA Pyrosequences
NSF 1062362 (June 1, 2011 – May 31, 2014)
PI: Sun, Co-PI: Mai
The University of Florida is awarded a grant to develop an integrated suite of computational tools and statistical methods that allow researchers to analyze tens of millions of 16S rRNA sequences for microbial community analysis and to extract biologically relevant patterns from massive sequence data. These tools will be made available to the research community as a web application. Microbes play an essential role in processes as diverse as human health and biogeochemical activities critical to life in all environments on earth. Complex microbial communities, however, remain poorly characterized. Currently available pyrosequencing technologies can determine nucleotide sequences of millions of individual 16S rRNA molecules in a matter of hours, opening new windows to probe the hidden microbial world. However, such large amounts of data overwhelm existing computational resources and analytic methods. An interdisciplinary research plan will be used to develop computational algorithms to overcome the current computational hurdles of large-scale 16S rRNA based analysis of microbial communities. Advanced computational techniques will be used, including parallel computing, online learning, graphical modeling, supervised and unsupervised learning, and dimensionality reduction. The specific aims include: (1) to develop computational algorithms for large-scale taxonomy independent analysis; (2) to develop a collection of statistical and computational methods for comparative community analysis, including discriminant analysis, topology analysis and microbial network analysis; (3) to establish a web application based on the proposed algorithms to provide researchers with a complete package of tools to perform comparative microbial community analysis. The analytical approaches developed in the project will enable the derivation of microbial community diversity, quantitative disease-associated microbial profiles, environment-microbe and microbe-microbe interactions, and will identify and quantify sequences from unclassified species. Many of the analytic methods that will be developed have not been traditionally used to analyze microbial communities. Hence, this work represents a major transformation of the bioinformatics methodology used for investigating microbial communities, and has the potential to significantly advance discovery and understanding of the hidden microbial world.
The results from this study will be disseminated through publications, web applications, workshops and open source projects. Multiple impacts are anticipated. The development of new computational approaches capable of efficiently handling tens of millions of sequences currently generated by the third-generation sequencers can greatly improve the utility of existing pipelines. Open source projects will invite researchers from other fields such as mathematics and statistics to join this project. The close interactions between computer scientists and biologists that we propose to develop will create new teaching and training opportunities, and spark new algorithmic research with direct utility to biologists. Currently, there is a shortage of researchers with a deep understanding of both computer science and molecular biology. This project will provide two graduate students and a postdoctoral fellow with an excellent opportunity to receive intensive training in both areas. Software and results of this project will be available at http://plaza.ufl.edu/sunyijun/DProject.htm
Metagenomic based discoveries of new viral and eukaryotic pathogens
Bill and Melinda Gates Foundation (January 1, 2011 – December 31, 2013)
Co-PI on UFL subcontract: Mai
In this study we are utilizing high throughput molecular tools to discover potentially novel diarrhea viruses in children in Mali, Kenya, India and China. Fecal samples are being collected from children with severe diarrhea as well as from controls matched by gender, age and location.
Microbiota in Preterm Infants and NEC
NIH 1 R01 HD059143-01 (December 17, 2008 – November 30, 2013)
PI: Neu, Co-PI: Mai
We propose the first comprehensive evaluation of the intestinal microbiota in premature infants at risk for necrotizing enterocolitis (NEC). The overall goals respond to objectives of the NIH Funding Opportunity Announcement, which include: 1. gaining a better understanding of the bacterial ecology of the infant intestine, 2. understanding how commensal organisms work to dampen inappropriate inflammatory responses of the enterocyte, 3. find methods to better identify prematurely born infants at highest risk of NEC in order to enter them into clinical trials of preventive therapy. Three hypotheses will be tested: Hypothesis 1. Infants who subsequently develop NEC establish a high risk microbiota that can be distinguished from a low risk pattern. This will be accomplished using unique newly developed molecular techniques; Hypothesis 2. High-risk- for- NEC stool microbiota patterns are associated with altered inflammatory mediator markers. This will be evaluated using correlations with plasma samples as well as buccal smears, which may offer the potential for determination of early biomarkers using a relatively non-invasive approach; Hypothesis 3. The high risk fecal microbiota and inflammatory patterns are associated with other morbidities such as hospital acquired sepsis, chronic lung disease and neurological injury. Here we will correlate intestinal microbial patterns and inflammatory marker patterns with non intestinal diseases associated with inflammatory processes in the neonate such hospital acquired sepsis, chronic lung disease and neurologic damage such as intracranial hemorrhage (IVH) and periventricular leukomalacia (PVL). This research will provide new information on the relationship between intestinal microbiota, gut derived systemic inflammation, NEC and other neonatal morbidities. Significance: At the completion of this project a relationship between NEC and stool microbiota will be found: This may consist of finding an individual causative pathogen or more likely, a quantitative or qualitative pattern that is consistently associated with the presence or absence of NEC. This will guide us in the choice of safe and effective preventative measures, such as choice of correct probiotic. In addition, finding inflammation-related markers using a non-invasive technique such as fecal analysis or tongue scraping to determine individuals at highest risk for the development of NEC will improve our chances of only using this preventative therapy for those individuals at highest risk for the disease PUBLIC HEALTH REVELANCE: The relevance of this research is that at the completion of this project a relationship between NEC, one of the most devastating diseases seen in premature infants, and stool microbes will be found: This may consist of finding an individual causative pathogen or more likely, a quantitative or qualitative pattern that is consistently associated with the presence or absence of NEC. This will guide us in the choice of safe and effective preventative measures and result in an enhanced ability to find those at highest risk by use of non-invasive techniques such as fecal analysis or tongue swabs, rather than the more invasive and painful blood based analysis.
Probiotics on immune and intestinal health in healthy older adults
Wakunaga of America Co., Ltd (March, 01, 2012 – November 07, 2013)
PI: Langkamp-Henken, Co-PI: Mai
Probiotics are live bacteria that can colonize the human gut and help to improve or maintain health. While intake of the probiotic is known to increase the amounts of the probiotic that can be recovered from feces, other effects on microbiota composition are less well established. We will determine how the daily intake of 5 billion lactic acid bacteria (L. gasseri) affects levels of lactic acid bacteria in stool samples as well as the overall microbiota.
Effects of almond consumption on gut microbiota
California Almond Board (February 25, 2010 – June 30, 2013)
Tree nuts are thought to benefit health, but corresponding mechanisms are not fully established. We investigate in a controlled feeding study if increased intake of almonds affects fecal microbiota composition.
Bacteriophage-based probiotic preparation for managing Shigella infections
DoD AMSRD STTR Phase II, Subcontract with Intralytix, Inc. (June 01, 2011 – May 31, 2013)
PI Academic Partner: Mai
During the studies supported by our Phase I STTR grant from the US Army, we began developing and testing a novel concept for a probiotic preparation based on naturally occurring bacteriophages as a way to condition the GI tract”s microflora gently and favorably. The preparation (tentatively designated”ShigActive”) is a bacteriophage cocktail that specifically targets Shigella spp. (significant diarrhea-causing pathogens that sicken ca. 165 million people/year, ca. 1.1 million of whom die). We have succesfully isolated and rigorously characterized a large number of phages lytic for Shigella, and we have developed a murine model in which the in vivo efficacy of our phage preparation can be evaluated. During this Phase II project, we propose to perform studies required to obtain regulatory approval of ShigActive. During these studies, we will also perform metagenomic analyses of the gut microbiota (using DGGE and state-of-the-art 454-pyrosequencing of 16S rRNA genes) of mice before and after their treatment with ShigActive, in order to (i) obtain additional safety data for the FDA, and (ii) compare rigorously the effect of ShigActive and ampicillin (an antibiotic commonly used to treat shigellosis) on the intestinal microflora of mice. The proposed project”s primary goal is to submit a New Dietary Ingredient (NDI) Notification to the FDA, whose approval will enable us to commence commercial sales of ShigActive.
Immune modulation by adding fiber to the diet
General Mills (September 01, 2009 – December 31, 2012)
PI: Langkamp-Henken, Co-PI: Mai
This study was designed to determine the effects of increasing whole grain intake on various health markers in young school children. The families of each child enrolled in the study received either whole grain or refined grain products to be prepared and consumed at home. An emphasis of our study is the correlation between whole grain intake and composition of the commensal gut microbiota.
Effects of galacto-oligosacharides on immune function
GTC (January 01, 2010 – December 31, 2012)
PI: Langkamp-Henken, Co-PI: Mai
Here we evaluate the effects of increased galacto-oligosacharides (GOS) intake on the number of days with cold symptoms as a marker of immune function. As GOS represents a dietary fiber that can increase certain ‘beneficial’ gut bacteria we are analyzing the effects of GOS on commensal gut microbiota.
Diet, Microflora and Colorectal Carcinogenesis in African Americans
MRSGT CCE-107301 (July 01, 2004 – December 31, 2010)
African Americans suffer from an increased burden of colorectal cancer. We hypothesized that differences in the diet of African Americans results in a different microbiota that can contribute to increased CRC risk. IN this study we discovered a bacteria signature, based on 16S rRNA sequences, that is predictive of the presence of polyps and thus CRC risk.
New technologies for detection of diarrhea pathogens in developing countries
Bill and Melinda Gates Foundation (October 30, 2007 – December 31, 2010)
Co-PI of subcontract: Mai
In this study we are utilizing high throughput molecular tools to discover potentially novel bacterial diarrhea pathogens in children in Mali, Kenya, The Gambia and Bangladesh. Fecal samples were collected from children with severe diarrhea as well as from controls matched by gender, age and location. In the large 16S rRNA sequence dataset generated from 3000 samples we detected multiple signature sequences that are suggestive of the presence of hitherto unknown pathogens.
Bacteriophage-based probiotic preparation for managing Shigella infections
DoD AMSRD STTR Phase I, Subcontract with Intralytix, Inc. (January 01, 2010 – December 31, 2010)
PI Academic Partner: Mai
Diarrhea remains a major public health challenge worldwide, and it also may significantly hamper US troop operations during combat and peacekeeping missions overseas. Antibiotics can be used to treat diarrheal diseases; however, some foodborne and waterborne bacterial pathogens are developing resistance against antibiotics, which limits their effectiveness. Also, antibiotics – because of their broad spectrum activity – may disturb/alter the GI tract’s normal and beneficial microflora, which may create additional health problems. Therefore, novel approaches are needed to help prevent and treat bacterial-elicited diarrhea among US troops, thereby improving their combat readiness and performance. Synbiotics (combinations of probiotics and prebiotics) may provide one such approach. However, one potentially useful probiotic intervention that has not received much attention in the past is to use bacteriophages to target “problem” bacterial species in the human GI tract. Bacteriophages are viruses that infect bacteria, are highly specific, and lyse their targeted bacteria, including antibiotic-resistant strains. Thus, synbiotic preparations containing (i) bacteriophages targeting specific diarrhea-causing bacteria, (ii) previously described, bacteria-based probiotics, and (iii) prebiotics may provide excellent protection against diarrheal illnesses among US troops and civilian populations. In this application, we propose to begin developing and testing a synbiotic preparation whose phage component is specifically active against Shigella spp., which are significant diarrhea-causing pathogens.
Effects of bacteriophage administration against Listeria monocytogenes
NIH SBIR Phase I (May 01, 2008 – April 30, 2009)
PI Academic Partner: Mai
In this grant we established an animal model for the study of the effects of bacteriophage therapy. Animals received various regimen of bacteriophage therapy and were then challenged with Listeria monocytogenes. Recovery of Listeria monocytogenes in feces as well as translocation to various anatomic sites was studied
Diet, Intestinal Microflora and Colorectal Cancer
M01RR016500, Sub-Project ID: 6949 (March 1, 2005 – February 29, 2008)
This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Central hypothesis: Diet defines and modulates the intestinal microflora, with both diet and intestinal microflora serving as modifiable risk factors for the occurrence of colon cancer. rationale: The majority (2/3 or more) of colon cancer cases appear to be associated with non-hereditary, environmental factors. However, few studies to date have incorporated comprehensive data on the contents of the human large bowel, despite the approximately 1011 bacteria per gram that constitute >55% of total stool solids. The microbial ecosystem is one of the most complex known, with several hundred species and subspecies. Studies conducted using traditional microbiologic techniques have suggested that patterns of fecal flora differ among groups from different geographic areas, who are consuming different types of diets, and who have different cancer risks. Additional studies suggest that there are a number of bacterial strains/forms in stool samples that are visible by microscopy but that do not grow on cultures i.e., they non-culturable using standard microbiologic media and techniques. The advent of molecular technology has provided us with tools that permit rapid, comprehensive analysis of bacterial communities including both the culturable and +non-culturable+ bacteria. Whereas DGGE allows for an efficient profiling of the intestinal microflora, 16S rDNA sequencing allows for both the semiquantitative enumeration of the most prevalent gut bacteria and the detection of hitherto unknown gut microbes. With such tools, we are in a position, for the first time, to look for linkages among diet, intestinal flora, and colon cancer. The proposed pilot study with 60 subjects is part of a larger study that will investigate these associations in a total of 500 subjects. The larger study is funded by an ACS mentored research scholar grant to the PI. The pilot study will allow us to investigate the proposed associations in more depth and optimize the analysis approach for the larger study. The results from the 60 subject pilot study will also form the basis for the resubmission of an ambitious PPG to the NCI and thus hopefully lead to additional extramural funding.
Diarrhea in Baltimore: Surveillance & Pathogen Detection
U01CI000296 (October 01, 2004 – September 30, 2007)
PI: Hirshon, Co-I: Mai
There are an estimated 76 million illnesses, 325,000 hospitalizations, and 5,000 deaths per year due to food-related pathogens in the United States. Of these food related illnesses, approximately 62 million are due to undetermined pathogens. Considering this burden of disease, there is a critical need to improve our understanding of the causes of foodborne illnesses. Thus, this proposal is in response to a request for proposals from the National Center for Infectious Diseases. As a continuation of our previous work in exploring the causes of unexplained diarrhea through prior grants and through the Emerging Infections Program Sentinel Site Grant (aka: FoodNet), our goals for this work fall into three areas. First, we will continue to use the most sophisticated, currently available methodologies to define the causes of infectious diarrhea in patients seeking medical attention; second, we will determine the prevalence of antibiotic resistant enteric bacteria in the community; and third we will employ advanced molecular techniques to screen for novel pathogens. This research will allow us to increase our understanding of the causes of foodborne illnesses. Additionally, it has the potential for translation into public health practice in order to decrease the significant morbidity and mortality caused by these diseases. Considering the depth of expertise available at the University of Maryland, especially as it relates to enteric pathogens, we have an ideal environment to conduct these studies. Our three specific aims are: 1) Identify causes of diarrhea from known enteric pathogens in pediatric and adult populations presenting to the emergency departments and clinics within the University of Maryland Medical System, 2) Undertake antibiotic susceptibility determinations on selected enteric isolates from individuals with and without complaints of diarrhea, 3) Identify new pathogens by utilizing novel molecular methods (specifically denaturing gradient gel electrophoresis (DGGE), fluorescent in situ hybridization (FISH), and microarrays). The combination of sophisticated microbiologic techniques, antibiotic resistance determinations, and molecular methods provides an innovative approach to discovering the causes of gastroenteritis. Through this work, we expect to not only better define the causes of diarrheal illnesses in the Baltimore Metropolitan area, but to identify other pathogens that contribute to foodborne illnesses.